Model selection for prognostic time-to-event gene signature discovery with applications in early breast cancer data

Model selection between competing models is a key consideration in the discovery of prognostic multigene signatures. The use of appropriate statistical performance measures as well as verification of biological significance of the signatures is imperative to maximise the chance of external validation of the generated signatures. Current approaches in time-to-event studies often use only a single measure of performance in model selection, such as logrank test p-values, or dichotomise the follow-up times at some phase of the study to facilitate signature discovery. In this study we improve the prognostic signature discovery process through the application of the multivariate partial Cox model combined with the concordance index, hazard ratio of predictions, independence from available clinical covariates and biological enrichment as measures of signature performance. The proposed framework was applied to discover prognostic multigene signatures from early breast cancer data. The partial Cox model combined with the multiple performance measures were used in both guiding the selection of the optimal panel of prognostic genes and prediction of risk within cross validation without dichotomising the follow-up times at any stage. The signatures were successfully externally cross validated in independent breast cancer datasets, yielding a hazard ratio of 2.55 [1.44, 4.51] for the top ranking signature.

The significance of combining VEGFA, FLT1, and KDR expressions in colon cancer patient prognosis and predicting response to bevacizumab

Targeting angiogenesis through inhibition of the vascular endothelial growth factor (VEGF) pathway has been successful in the treatment of late stage colorectal cancer. However, not all patients benefit from inhibition of VEGF. Ras status is a powerful biomarker for response to anti-epidermal growth factor receptor therapy; however, an appropriate biomarker for response to anti-VEGF therapy is yet to be identified. VEGF and its receptors, FLT1 and KDR, play a crucial role in colon cancer progression; individually, these factors have been shown to be prognostic in colon cancer; however, expression of none of these factors alone was predictive of tumor response to anti-VEGF therapy. In the present study, we analyzed the expression levels of VEGFA, FLT1, and KDR in two independent colon cancer datasets and found that high expression levels of all three factors afforded a very poor prognosis. The observation was further confirmed in another independent colon cancer dataset, wherein high levels of expression of this three-gene signature was predictive of poor prognosis in patients with proficient mismatch repair a wild-type KRas status, or mutant p53 status. Most importantly, this signature also predicted tumor response to bevacizumab, an antibody targeting VEGFA, in a cohort of bevacizumab-treated patients. Since bevacizumab has been proven to be an important drug in the treatment of advanced stage colon cancer, our results suggest that the three-gene signature approach is valuable in terms of its prognostic value, and that it should be further evaluated in a prospective clinical trial to investigate its predictive value to anti-VEGF treatment.

The prognostic and therapeutic value of EpHA2 in early colorectal cancer (CRC).

Background: EpHA2 is a 130 kD transmembrane glycoprotein belonging to ephrin receptor subfamily and involved in angiogenesis/tumour neovascularisation. High EpHA2 mRNA level has recently been implicated in cetuximab resistance. Previously, we found high EpHA2 levels in a panel of invasive colorectal cancer (CRC) cells, which was associated with high levels of stem-cell marker CD44. Our aim was to investigate the prognostic value of EpHA2 and subsequently correlate expression levels to known clinico-pathological variables in early stage CRC. Methods: Tissue samples from 509 CRC patients were analysed. EpHA2 expression was measured using IHC. Kaplan-Meier graphs were used. Univariate and multivariate analyses employed Cox Proportional Hazards Ratio (HR) method. A backward selection method (Akaike’s information criterion) was used to determine a refined multivariate model. Results: EpHA2 was highly expressed in CRC adenocarcinoma compared to matched normal colon tissue. In support of our preclinical invasive models, strong correlation was found between EpHA2 expression and CD44 and Lgr5 staining (p<0.001). In addition, high EpHA2 expression significantly correlated with vascular invasion (p=0.03).HR for OS for stage II/III patients with high EpHA2 expression was 1.69 (95%CI: 1.164-2.439; p=0.003). When stage II/III was broken down into individual stages, there was significant correlation between high EpHA2 expression and poor 5-years OS in stage II patients (HR: 2.18; 95%CI: 1.28-3.71; p=0.005).HR in the stage III group showed a trend to statistical significance (HR: 1.48; 95%CI=0.87-2.51; p=0.05). In both univariate and multivariate analyses of stage II patients, high EpHA2 expression was the only significant factor and was retained in the final multivariate model. Higher levels of EpHA2 were noted in our RAS and BRAF mutant CRC cells, and silencing EpHA2 resulted in significant decreases in migration/invasion in parental and invasive CRC sublines. Correlation between KRAS/NRAS/BRAFmutational status and EpHA2 expression in clinical samples is ongoing. Conclusions: Taken together, our study is the first to indicate that EpHA2 expression is a predictor of poor clinical outcome and a potential novel target in early stage CRC.

Logistic regression analysis of potential prognostic factors for pulmonary thromboembolism

Objective: To identify potential prognostic factors for pulmonary thromboembolism (PTE), establishing a mathematical model to predict the risk for fatal PTE and nonfatal PTE.Method: the reports on 4,813 consecutive autopsies performed from 1979 to 1998 in a Brazilian tertiary referral medical school were reviewed for a retrospective study. From the medical records and autopsy reports of the 512 patients found with macroscopically and/or microscopically,documented PTE, data on demographics, underlying diseases, and probable PTE site of origin were gathered and studied by multiple logistic regression. Thereafter, the jackknife method, a statistical cross-validation technique that uses the original study patients to validate a clinical prediction rule, was performed.Results: the autopsy rate was 50.2%, and PTE prevalence was 10.6%. In 212 cases, PTE was the main cause of death (fatal PTE). The independent variables selected by the regression significance criteria that were more likely to be associated with fatal PTE were age (odds ratio [OR], 1.02; 95% confidence interval [CI], 1.00 to 1.03), trauma (OR, 8.5; 95% CI, 2.20 to 32.81), right-sided cardiac thrombi (OR, 1.96; 95% CI, 1.02 to 3.77), pelvic vein thrombi (OR, 3.46; 95% CI, 1.19 to 10.05); those most likely to be associated with nonfatal PTE were systemic arterial hypertension (OR, 0.51; 95% CI, 0.33 to 0.80), pneumonia (OR, 0.46; 95% CI, 0.30 to 0.71), and sepsis (OR, 0.16; 95% CI, 0.06 to 0.40). The results obtained from the application of the equation in the 512 cases studied using logistic regression analysis suggest the range in which logit p > 0.336 favors the occurrence of fatal PTE, logit p < - 1.142 favors nonfatal PTE, and logit P with intermediate values is not conclusive. The cross-validation prediction misclassification rate was 25.6%, meaning that the prediction equation correctly classified the majority of the cases (74.4%).Conclusions: Although the usefulness of this method in everyday medical practice needs to be confirmed by a prospective study, for the time being our results suggest that concerning prevention, diagnosis, and treatment of PTE, strict attention should be given to those patients presenting the variables that are significant in the logistic regression model.

Immunocytochemical study of Ki-67 as a prognostic marker in canine mammary neoplasia

Background: Canine mammary tumors are challenging for clinicians and pathologists because of complex histologic classification, low specificity of cytologic diagnosis, and unpredictable biological behavior. In histologic specimens, expression of tumor proliferation marker Ki-67, a nuclear nonhistone protein, has been shown to have prognostic value for canine mammary tumors and to correlate with malignancy and low survival rates. Objective: The objective of this study was to measure the proliferation index of canine mammary tumors by immunochemical detection of Ki-67 in cytologic specimens and to determine its relationship to clinical and pathologic variables and patient outcome. Methods: Spontaneous mammary tumors from 31 female dogs were surgically excised. Imprint specimens for cytologic evaluation were wet-fixed in ethanol; histologic specimens were prepared routinely. Immunostaining was performed with the PH 177 monoclonal antibody against Ki-67; proliferation index was graded from negative to +++. Dogs were followed for 18 months. Multivariate logistic regression analysis was used to determine correlations between immunocytochemical results, tumor and clinical variables, and patient outcome. Results: Ki-67 proliferation indices in cytologic specimens were significantly lower for nonmalignant tumors than for malignant tumors. High index values of Ki-67 were positively correlated with metastasis, death from neoplasia, low disease-free survival rates, and low overall survival rate. With the exception of 4 specimens for which cellularity was insufficient, positive expression of Ki-67 in cytologic specimens correlated with that of histologic specimens. Conclusions: The prognostic value of the Ki-67 index in canine mammary tumors by using wet-fixed cytology imprint specimens was similar to that observed previously for histologic specimens. Immunocytochemical detection of Ki-67 could improve the accuracy and value of cytology by providing safe and rapid information about malignancy and patient outcome. © 2004 American Society for Veterinary Clinical Pathology.

Clinicopathological significance of ERCC1 expression in breast cancer

The excision repair cross-complementation 1 (ERCC1) enzyme plays an essential role in the nucleotide excision repair pathway and is associated with resistance to platinum-based chemotherapy in different types of cancer. The aim of the present study was to evaluate the clinicopathological significance of ERCC1 expression in breast cancer patients. We analyzed the immunohistochemical expression of ERCC1 in a tissue microarray from 135 primary breast carcinomas and correlated the immunohistochemical findings with clinicopathological factors and outcome data. ERCC1 expression analysis was available for 109 cases. In this group, 58 (53.2%) were positive for ERCC1. ERCC1-positive expression was correlated with smaller tumor size (P=0.007) and with positivity for estrogen receptor (P=0.040), but no correlation was found with other clinicopathological features. Although not statistically significant, triple negative breast cancers were more frequently negative for ERCC1 (61.5% of the cases) compared to the non-triple negative breast cancer cases (41.5%). In conclusion, ERCC1 expression correlated significantly with favorable prognostic factors, such as smaller tumor size and ER-positivity, suggesting a possible role for ERCC1 as a predictive and/or prognostic marker in breast cancer. © 2013 Elsevier GmbH.

The density of metastatic lymph node as prognostic factor in squamous cell carcinoma of the tongue and floor of the mouth

The presence of metastatic lymph nodes is a relevant prognostic factor in oral cancer. Objective: This paper aims to assess metastatic lymph node density (pN+) in patients with tongue and floor-of-mouth squamous cell carcinoma (SCC) and the association of this parameter with disease-free survival (DFS). Materials and Methods: A group of 182 patients seen between 1985 and 2007 was included, 169 of which were males. Five were on stage I, 35 on stage II, 56 on stage III, and 85 on stage IV. Median values were considered in lymph node density assessment, and the Kaplan-Meier curve was used to evaluate DFS; survival differences within the group were elicited through the log-rank test. Results: An average 3.2 metastatic lymph nodes were excised from the patients in the group. Density ranged from 0.009 to 0.4, with a mean value of 0.09. Five-year DFS rates were of 44% and 28% for the groups with lymph node densities below and above the median respectively (p = 0.006). Two-year local/regional control was achieved for 71% and 49% for the patients below and above the median density respectively (p = 0.01). In terms of pN staging, local/regional control was achieved in 70% and 54% of pN1 and pN2 patients respectively, albeit without statistical significance (0.20%). Conclusion: Lymph node density may be used as a prognostic indicator for tongue and floor-ofmouth SCC.

Validity and clinical significance of biomechanical testing of implant/bone interface

Purpose: The aim of this paper was to review the clinical literature on the Resonance frequency analysis (RFA) and Periotest techniques in order to assess the validity and prognostic value of each technique to detect implants at risk for failure. Material and methods: A search was made using the PubMed database to find clinical studies using the RFA and/or Periotest techniques. Results: A limited number of clinical reports were found. No randomized-controlled clinical trials or prospective cohort studies could be found for validity testing of the techniques. Consequently, only a narrative review was prepared to cover general aspects of the techniques, factors influencing measurements and the clinical relevance of the techniques. Conclusions: Factors such as bone density, upper or lower jaw, abutment length and supracrestal implant length seem to influence both RFA and Periotest measurements. Data suggest that high RFA and low Periotest values indicate successfully integrated implants and that low/decreasing RFA and high/increasing Periotest values may be signs of ongoing disintegration and/or marginal bone loss. However, single readings using any of the techniques are of limited clinical value. The prognostic value of the RFA and Periotest techniques in predicting loss of implant stability has yet to be established in prospective clinical studies. To cite this article: Aparicio C, Lang N P, Rangert B. Validity and clinical significance of biomechanical testing of implant/bone interface. Clin. Oral Imp. Res., 17 (Suppl. 2), 2006; 2-7.

Clinical significance of cell cycle- and apoptosis-related markers in biliary tract cancer: a tissue microarray-based approach revealing a distinctive immunophenotype for intrahepatic and extrahepatic cholangiocarcinomas

Cholangiocarcinoma is the second most common malignant tumor of the liver. We analyzed, immunohistochemically, the significance of cell cycle- and apoptosis-related markers in 128 cholangiocarcinomas (42 intrahepatic, 70 extrahepatic, and 16 gallbladder carcinomas) combined in a tissue microarray. Follow-up was available for 57 patients (44.5%). In comparison with normal tissue (29 specimens), cholangiocarcinomas expressed significantly more frequently p53, bcl-2, bax, and COX-2 (P.05 <). Intrahepatic tumors were significantly more frequently bcl-2+ and p16+, whereas extrahepatic tumors were more often p53+ (P < .05). Loss of p16 expression was associated with reduced survival of patients. Our data show that p53, bcl-2, bax, and COX-2 have an important role in the pathogenesis of cholangiocarcinomas. The differential expression of p16, bcl-2, and p53 between intrahepatic and extrahepatic tumors demonstrates that there are location-related differences in the phenotype and the genetic profiles of these tumors. Moreover, p16 was identified as an important prognostic marker in cholangiocarcinomas.

Septic peritonitis from pyloric and non-pyloric gastrointestinal perforation: prognostic factors in 44 dogs and 11 cats

OBJECTIVES To identify potential prognostic factors affecting outcome in septic peritonitis caused by gastrointestinal perforation in dogs and cats. METHODS A retrospective study. Animals operated on for septic peritonitis because of gastrointestinal perforation were evaluated. Risk factors assessed included age, duration of clinical signs, recent prior abdominal surgery, recent prior anti-inflammatory drug administration, placement of a closed-suction drain and location of perforation. RESULTS Fifty-five animals (44 dogs and 11 cats) were included. The overall mortality was 63·6%. No association was found between age, duration of clinical signs or prior abdominal surgery and outcome. Animals with a history of prior anti-inflammatory drugs were significantly (P=0·0011) more likely to have perforation of the pylorus (73·3%). No significant difference in outcome was found between animals treated with closed-suction drains and those treated with primary closure or between pyloric perforation and perforation at other gastrointestinal sites. CLINICAL SIGNIFICANCE Administration of anti-inflammatory drugs in dogs and cats is a significant risk factor for pyloric perforation. Pyloric perforation was not associated with a poorer outcome than perforation at other gastrointestinal sites. Placement of a closed suction drain did not improve outcome compared to primary closure.

Clinical Significance of NOTCH1 and NOTCH2 Expression in Gastric Carcinomas: An Immunohistochemical Study.

BACKGROUND NOTCH signaling can exert oncogenic or tumor suppressive functions and can contribute to chemotherapy resistance in cancer. In this study, we aimed to clarify the clinicopathological significance and the prognostic and predictive value of NOTCH1 and NOTCH2 expression in gastric cancer (GC). METHODS NOTCH1 and NOTCH2 expression was determined immunohistochemically in 142 primarily resected GCs using tissue microarrays and in 84 pretherapeutic biopsies from patients treated by neoadjuvant chemotherapy. The results were correlated with survival, response to therapy, and clinico-pathological features. RESULTS Primarily resected patients with NOTCH1-negative tumors demonstrated worse survival. High NOTCH1 expression was associated with early-stage tumors and with significantly increased survival in this subgroup. Higher NOTCH2 expression was associated with early-stage and intestinal-type tumors and with better survival in the subgroup of intestinal-type tumors. In pretherapeutic biopsies, higher NOTCH1 and NOTCH2 expression was more frequent in non-responding patients, but these differences were statistically not significant. CONCLUSION Our findings suggested that, in particular, NOTCH1 expression indicated good prognosis in GC. The close relationship of high NOTCH1 and NOTCH2 expression with early tumor stages may indicate a tumor-suppressive role of NOTCH signaling in GC. The role of NOTCH1 and NOTCH2 in neoadjuvantly treated GC is limited.

Prognostic and diagnostic value of EEG signal coupling measures in coma.

OBJECTIVE Our aim was to assess the diagnostic and predictive value of several quantitative EEG (qEEG) analysis methods in comatose patients. METHODS In 79 patients, coupling between EEG signals on the left-right (inter-hemispheric) axis and on the anterior-posterior (intra-hemispheric) axis was measured with four synchronization measures: relative delta power asymmetry, cross-correlation, symbolic mutual information and transfer entropy directionality. Results were compared with etiology of coma and clinical outcome. Using cross-validation, the predictive value of measure combinations was assessed with a Bayes classifier with mixture of Gaussians. RESULTS Five of eight measures showed a statistically significant difference between patients grouped according to outcome; one measure revealed differences in patients grouped according to the etiology. Interestingly, a high level of synchrony between the left and right hemisphere was associated with mortality on intensive care unit, whereas higher synchrony between anterior and posterior brain regions was associated with survival. The combination with the best predictive value reached an area-under the curve of 0.875 (for patients with post anoxic encephalopathy: 0.946). CONCLUSIONS EEG synchronization measures can contribute to clinical assessment, and provide new approaches for understanding the pathophysiology of coma. SIGNIFICANCE Prognostication in coma remains a challenging task. qEEG could improve current multi-modal approaches.

Analysis of prognostic factors for the development of metastases and survival in renal cell carcinoma patients

Introduction and objective. A number of prognostic factors have been reported for predicting survival in patients with renal cell carcinoma. Yet few studies have analyzed the effects of those factors at different stages of the disease process. In this study, different stages of disease progression starting from nephrectomy to metastasis, from metastasis to death, and from evaluation to death were evaluated. ^ Methods. In this retrospective follow-up study, records of 97 deceased renal cell carcinoma (RCC) patients were reviewed between September 2006 to October 2006. Patients with TNM Stage IV disease before nephrectomy or with cancer diagnoses other than RCC were excluded leaving 64 records for analysis. Patient TNM staging, Furhman Grade, age, tumor size, tumor volume, histology and patient gender were analyzed in relation to time to metastases. Time from nephrectomy to metastasis, TNM staging, Furhman Grade, age, tumor size, tumor volume, histology and patient gender were tested for significance in relation to time from metastases to death. Finally, analysis of laboratory values at time of evaluation, Eastern Cooperative Oncology Group performance status (ECOG), UCLA Integrated Staging System (UISS), time from nephrectomy to metastasis, TNM staging, Furhman Grade, age, tumor size, tumor volume, histology and patient gender were tested for significance in relation to time from evaluation to death. Linear regression and Cox Proportional Hazard (univariate and multivariate) was used for testing significance. Kaplan-Meier Log-Rank test was used to detect any significance between groups at various endpoints. ^ Results. Compared to negative lymph nodes at time of nephrectomy, a single positive lymph node had significantly shorter time to metastasis (p<0.0001). Compared to other histological types, clear cell histology had significant metastasis free survival (p=0.003). Clear cell histology compared to other types (p=0.0002 univariate, p=0.038 multivariate) and time to metastasis with log conversion (p=0.028) significantly affected time from metastasis to death. A greater than one year and greater than two year metastasis free interval, compared to patients that had metastasis before one and two years, had statistically significant survival benefit (p=0.004 and p=0.0318). Time from evaluation to death was affected by greater than one year metastasis free interval (p=0.0459), alcohol consumption (p=0.044), LDH (p=0.006), ECOG performance status (p<0.001), and hemoglobin level (p=0.0092). The UISS risk stratified the patient population in a statistically significant manner for survival (p=0.001). No other factors were found to be significant. ^ Conclusion. Clear cell histology is predictive for both time to metastasis and metastasis to death. Nodal status at time of nephrectomy may predict risk of metastasis. The time interval to metastasis significantly predicts time from metastasis to death and time from evaluation to death. ECOG performance status, and hemoglobin levels predicts survival outcome at evaluation. Finally, UISS appropriately stratifies risk in our population. ^

Prognostic value of the diagnostic criteria distinguishing endometrial stromal sarcoma, low grade from undifferentiated endometrial sarcoma, 2 entities within the invasive endometrial stromal neoplasia family

The World Health Organization (WHO 2003) recognizes 3 endometrial stromal neoplasms: noninvasive endometrial stromal nodule and the 2 invasive neoplasms, endometrial stromal sarcoma (ESS), low grade and undifferentiated endometrial sarcoma (UES). It is important to note that the WHO 2003 does not define moderate atypia (an important differentiating diagnostic criterion for ESS, low grade and UES), nor does it discuss its significance. Moreover, studies on reproducibility and additional prognostic value of other diagnostic features in large are lacking. Using strict definitions, we analyzed the agreement between routine and expert-review necrosis and nuclear atypia in 91 invasive endometrial stromal neoplasias (IESN). The overall 5-year and 10-year recurrence-free survival rate estimates of the 91 IESN patients were 82% and 75%, respectively. Necrosis was well reproducible, and nuclear atypia was reasonably well reproducible. The 10-year recurrence-free survival rates for necrosis absent/inconspicuous versus prominent were 89% and 45% (P<0.001) and those for review-confirmed none/mild, moderate, severe atypia were 90%, 30%, and <20% (P<0.00001). Therefore, cases with moderate/severe atypia should be grouped together. Nuclear atypia and necrosis had independent prognostic values (Cox regression). Once these features were taken into account, no other feature had an independent additional prognostic value, including mitotic count. Using "none/mild atypia, necrosis absent/inconspicuous" as ESS, low grade versus "moderate/severe atypia present or necrosis present" as UES resulted in 68 ESS, low grade and 23 UES cases with disease-specific overall mortality-free survival of 99% versus 48% (P<0.00001, hazard ratio=45.4). When strictly defined microscopic criteria are used, the WHO 2003 diagnoses of ESS, low grade and UES are well reproducible and prognostically strong. © 2012 International Society of Gynecological Pathologists.

PROGNOSTIC FACTORS PREDICTING FUNCTIONAL OUTCOME AT FOUR MONTHS FOLLOWING ACUTE ANKLE SPRAIN

PROGNOSTIC FACTORS PREDICTING FUNCTIONAL OUTCOME AT FOUR MONTHS FOLLOWING ACUTE ANKLE SPRAINBleakley C.M.1, O'Connor S.R.1, Tully M.A.2, Rocke L.G.3, MacAuley D.C.1, Bradbury I.4, Keegan S.4, McDonough S.M.11University of Ulster, Health & Rehabilitation Sciences Research Institute, Newtownabbey, United Kingdom, 2Queen's University, UKCRC Centre of Excellence for Public Health (NI), Belfast, United Kingdom, 3Royal Victoria Hospital, Department of Emergency Medicine, Belfast, United Kingdom, 4Frontier Science (Scotland), Kincraig, Inverness-shire, United KingdomPurpose: To identify clinically relevant factors assessed following acute ankle sprain that predict functional recovery at four months post-injury.Relevance: Ankle sprains are one of the most common musculoskeletal injuries with an estimated 5000 new cases occurring each day in the United Kingdom. In the acute phase, ankle sprains may be associated with pain and loss of function. In the longer-term there is a risk of residual problems including chronic pain or reinjury. Few studies have sought to examine factors associated with a poor long-term prognosis.Participants: 101 patients (Age: Mean (SD) 25.9 (7.9) years; Body Mass Index (BMI): 25.3 (3.5) kg/m2) with an acute grade 1 or 2 ankle sprain attending an accident and emergency department or sports injury clinic. Exclusion criteria included complete (grade 3) rupture of the ankle ligament complex, bony ankle injury or multiple injuries.Methods: Participants were allocated as part of a randomised controlled trial to an accelerated intervention incorporating intermittent ice and early therapeutic exercise or a standard protection, rest, ice, compression, and elevation intervention for one week. Treatment was then standardised in both groups and consisted of ankle rehabilitation exercises focusing on muscle strengthening, neuromuscular training, and sports specific functional exercises for a period of approximately four to six weeks. On initial assessment age, gender, mechanism of injury, presence of an audible pop or snap and the presence of contact during the injury were recorded. The following factors were also recorded at baseline and at one and four weeks post-injury: weight-bearing dorsi-flexion test, lateral hop test, presence of medial pain on palpation and a positive impingement sign. Functional status was assessed using the Karlsson score at baseline, at week four and at four months. Reinjury rates were recorded throughout the intervention phase and at four months.Analysis: A mixed between-within subjects analysis of variance (ANOVA) was used to determine the effect of each factor on functional status at week four and at four months. Significance was set at a Bonferroni adjusted level of 0.0125 (0.05/4).Results: Eighty-five participants (84%) were available at final follow-up assessment. Pain on weight-bearing dorsi-flexion and lateral hop tests at week four were both associated with a lower functional score at four months post-injury (P = 0.011 and P = 0.001). No other significant interactions were observed at any other timepoint (baseline or week one). There were only two reinjuries within the four month follow-up period with a further two reported at approximately six months post-injury. We were therefore unable to determine whether any factors were associated with an increased risk of reinjury.Conclusions: Potential prognostic factors on initial or early examination after acute ankle sprain did not help predict functional recovery at four months post-injury. However, pain on weight-bearing dorsi-flexion and lateral hop tests observed at four weeks were associated with a slower rate of recovery.Implications: Some clinical tests may help identify patients at risk of poor functional recovery after acute ankle sprain. However, further work is required to examine factors which may be predictive on initial assessment.Key-words: 1. Prognostic factors 2. Recovery 3. Ankle sprainFunding acknowledgements: Physiotherapy Research Foundation, Chartered Society of Physiotherapy, Strategic Priority Fund; Department of Employment and Learning, Northern Ireland.Ethics approval: Office for Research Ethics Committee (UK).