Validation of absolutely quantitated Ki67 and cyclinD1 protein levels for prognosis of Luminal-like breast cancer patients.

AIMS: To translate a clinical research finding into daily clinical practice requires well-controlled clinical trials. We have demonstrated the usage of absolute quantitation of Ki67 and cyclinD1 protein levels to improve prognosis of Luminal-like patients based on overall survival (OS) analysis of a cohort of 155 breast cancer specimens (cohort 1). However, this finding is considered the D level of evidence (LOE) to require subsequent validation before it may be used in daily clinical practice. To set the stage for future clinical trials, our findings were validated through OS analysis of an independent cohort (cohort 2) of 173 Luminal-like patients. METHODS: Both Ki67 and cyclinD1 levels were measured absolutely and quantitatively using the Quantitative Dot Blot (QDB) method in cohort 2. The proposed cutoffs for both biomarkers from cohort 1 were re-evaluated in cohort 2 and in the merged cohort of 1 and 2, respectively, through univariate, multivariate and Kaplan-Meier survival analysis. RESULTS: The proposed cutoffs of 2.31 nmol/g for Ki67 and 0.44 µmol/g for cyclinD1 were validated as effective cutoffs in cohort 2 and the merged cohort through OS analysis. The combined use of both biomarkers allowed us to identify patients with both biomarker levels below the cutoffs (59.3%) with10-year survival probability (SP) of 89%, in comparison to those above the cutoffs (8.3%) with 8 year SP of 28% through OS analysis in the merged cohort. CONCLUSIONS: This study validated our findings that absolute quantitation of Ki67 and cyclinD1 allows effective subtyping of luminal-like patients. It sets the stage for prospective or prospective-retrospective clinical studies.

New clues for early management of maxillary impacted central incisors based on 3-dimensional reconstructed models.

INTRODUCTION: The objective of this study was to provide new clues for the prevention and early management of root dilacerations in impacted maxillary central incisors. METHODS: Cone-beam computed tomography images of 108 patients with unilateral impacted maxillary central incisors were obtained and reconstructed into 3-dimensional models. Crown direction, crown height, root length, bone thickness, and position and angle of root dilaceration were measured in the sagittal-view sections. K-value, defined as the ratio between the available length of the direct root and the ideal length of the direct root, was proposed, and the relationships between K-values with root dilacerations were studied. Root development of the contralateral erupted maxillary incisor was also assessed. Independent t test, chi-square test, and 1-way analysis of variance were used for data analysis. RESULTS: Root dilacerations occurred when the K-values were 0.16 to 0.19 (palatal impaction), 0.25 to 0.53 (labial impaction), and 0.69 to 0.75 (nasal impaction). The position and angle of root dilacerations were different among nasal, labial, and palatal impactions (P <0.01). K-values and positions of root dilacerations among nasally, labially, and palatally impacted incisors were in descending order, respectively. Retarded root formation was noted in the impacted incisors compared with the contralateral incisors (P <0.001). CONCLUSIONS: Nasal, labial, and palatal impacted incisors had different patterns of root dilacerations. Analyses of crown direction and K-value may aid in evaluating root dilacerations at early dental ages and facilitating early intervention of impacted incisors.

Combined Use of cyclinD1 and Ki67 for Prognosis of Luminal-Like Breast Cancer Patients.

BACKGROUND: Ki67 is a biomarker of proliferation to be used in immunohistochemistry (IHC)-based surrogate assay to determine the necessity of cytotoxic therapy for Luminal-like breast cancer patients. cyclinD1 is another frequently used biomarker of proliferation. A retrospective study was performed here to investigate if these two biomarkers may be combined to improve the prognosis of Luminal-like patients. METHODS: Both Ki67 and cyclinD1 protein levels were measured absolutely and quantitatively using Quantitative Dot Blot method in 143 Luminal-like specimens. Optimized cutoffs for these two biomarkers were developed to evaluate their prognostic roles using Kaplan-Meier overall survival (OS) analysis. RESULTS: cyclinD1 was found as an independent prognostic factor from Ki67 in univariate and multivariate OS analyses. At optimized cutoffs (cyclinD1 at 0.44 µmol/g and Ki67 at 2.31 nmol/g), the subgroup with both biomarkers below the cutoffs (n = 65) had 10-year survival probability at 90% in comparison to those with both biomarkers above the cutoffs (n = 18) with 8-year survival probability at 26% (log-rank test, p <0.0001). This finding was used to modify the surrogate assay using IHC-based cyclinD1 scores, with p-value decreased from 0.031 to 0.00061 or from 0.1 to 0.02, when the Ki67 score of 14 or 20% was used as cutoff, respectively, in the surrogate assay. CONCLUSION: The current study supports the prospective investigation of cyclinD1 relevance in the clinic.

Improving Prognosis of Surrogate Assay for Breast Cancer Patients by Absolute Quantitation of Ki67 Protein Levels Using Quantitative Dot Blot (QDB) Method.

BACKGROUND: Immunohistochemistry (IHC)-based surrogate assay is the prevailing method in daily clinical practice to determine the necessity of chemotherapy for Luminal-like breast cancer patients worldwide. It relies on Ki67 scores to separate Luminal A-like from Luminal B-like breast cancer subtypes. Yet, IHC-based Ki67 assessment is known to be plagued with subjectivity and inconsistency to undermine the performance of the surrogate assay. A novel method needs to be explored to improve the clinical utility of Ki67 in daily clinical practice. MATERIALS AND METHODS: The Ki67 protein levels in a cohort of 253 specimens were assessed with IHC and quantitative dot blot (QDB) methods, respectively, and used to assign these specimens into Luminal A-like and Luminal B-like subtypes accordingly. Their performances were compared with the Kaplan-Meier, univariate, and multivariate survival analyses of the overall survival (OS) of Luminal-like patients. RESULTS: The surrogate assay based on absolutely quantitated Ki67 levels (cutoff at 2.31 nmol/g) subtyped the Luminal-like patients more effectively than that based on Ki67 scores (cutoff at 14%) (Log rank test, p = 0.00052 vs. p = 0.031). It is also correlated better with OS in multivariate survival analysis [hazard ratio (HR) at 6.89 (95% CI: 2.66-17.84, p = 0.0001) vs. 2.14 (95% CI: 0.89-5.11, p = 0.087)]. CONCLUSIONS: Our study showed that the performance of the surrogate assay may be improved significantly by measuring Ki67 levels absolutely, quantitatively, and objectively using the QDB method.

Botulinum toxin A alleviates orofacial nociception induced by orthodontic tooth movement through nociceptin/orphanin-FQ pathway in rats.

OBJECTIVES: To investigate the effect and mechanism of botulinum neurotoxin type A (BoNT/A) in the modulation of orofacial nociception induced by orthodontic tooth movement in rats. METHODS: An orofacial nociception model was established in male Sprague-Dawley rats by ligating closed-coil springs between incisors and ipsilateral molars. There were two group sets of animals. For the first group set, 120 rats were randomly divided into four groups: no-force group (n = 30), force + saline group (n = 30), force + low dose BoNT/A group (1U/6 µL, n = 30), and force + high dose BoNT/A group (1U/6 µL, n = 30). BoNT/A and saline were injected into periodontal ligament to explore the nociceptive effect of BoNT/A. Ipsilateral trigeminal ganglia (TG) were harvested for detecting the expression levels of nociceptin/orphanin-FQ (N/OFQ). For the second group set, 36 rats were randomly divided into three force groups: BoNT/A + saline group (n = 12), BoNT/A + UFP-101 group (n = 12), and saline + UFP-101 group (n = 12). A potent N/OFQ receptor (NOP) antagonist (UFP-101) was used to examine the role of N/OFQ in BoNT/A-induced antinociception. Tooth-movement nociception level of all groups was evaluated by bite force and rat grimace scale (RGS) at baseline, day 1, day 3, day 5, day 7, day 14. RESULTS: The behavioral assessments showed the orofacial nociception level in the force + low dose BoNT/A group and force + high dose BoNT/A group were lower than that in the force + saline group. No significant difference was observed in orofacial nociception among no-force group, force + low dose and force + high dose group. The expression levels of N/OFQ in TG were elevated from day 1 and maintained a high level, presenting in descending order among the force + high dose, force + low dose, force + saline and no-force group, respectively. The nociception level of the BoNT/A + UFP-101 group was higher than that of the BoNT/A + saline group. No significant difference was observed between the BoNT/A + UFP-101 group and the saline + UFP-101 group. CONCLUSIONS: BoNT/A can exert an antinociceptive effect on orofacial nociception induced by tooth movement by stimulating the expression of N/OFQ in TG.