Kathy Han

PKB/Akt phosphorylates p27, impairs nuclear import of p27 and opposes p27-mediated G1 arrest.

Mechanisms linking mitogenic and growth inhibitory cytokine signaling and the cell cycle have not been fully elucidated in either cancer or in normal cells. Here we show that activation of protein kinase B (PKB)/Akt, contributes to resistance to antiproliferative signals and breast cancer progression in part by impairing the nuclear import and action of p27. Akt transfection caused cytoplasmic p27 accumulation and resistance to cytokine-mediated G1 arrest. The nuclear localization signal of p27 contains an Akt consensus site at threonine 157, and p27 phosphorylation by Akt impaired its nuclear import in vitro. Akt phosphorylated wild-type p27 but not p27T157A. In cells transfected with constitutively active AktT308DS473D (PKBDD), p27WT mislocalized to the cytoplasm, but p27T157A was nuclear. In cells with activated Akt, p27WT failed to cause G1 arrest, while the antiproliferative effect of p27T157A was not impaired. Cytoplasmic p27 was seen in 41% (52 of 128) of primary human breast cancers in conjunction with Akt activation and was correlated with a poor patient prognosis. Thus, we show a novel mechanism whereby Akt impairs p27 function that is associated with an aggressive phenotype in human breast cancer.NOTE: In the version of the article initially published online, the abstract contained one extraneous sentence. This error has been corrected in the HTML and PDF versions. The abstract will appear correctly in the forthcoming print issue.

Phosphorylation of p27Kip1 regulates assembly and activation of cyclin D1-Cdk4

ABSTRACT p27 mediates Cdk2 inhibition and is also found in cyclin D1-Cdk4 complexes. The present data support a role for p27 in the assembly of D-type cyclin-Cdk complexes and indicate that both cyclin D1-Cdk4-p27 assembly and kinase activation are regulated by p27 phosphorylation. Prior work showed that p27 can be phosphorylated by protein kinase B/Akt (PKB/Akt) at T157 and T198. Here we show that PKB activation and the appearance of p27pT157 and p27pT198 precede p27-cyclin D1-Cdk4 assembly in early G1. PI3K/PKB inhibition rapidly reduced p27pT157 and p27pT198 and dissociated cellular p27-cyclin D1-Cdk4. Mutant p27 allele products lacking phosphorylation at T157 and T198 bound poorly to cellular cyclin D1 and Cdk4. Cellular p27pT157 and p27pT198 coprecipitated with Cdk4 but were not detected in Cdk2 complexes. The addition of p27 to recombinant cyclin D1 and Cdk4 led to cyclin D1-Cdk4-p27 complex formation in vitro. p27 phosphorylation by PKB increased p27-cyclin D1-Cdk4 assembly in vitro but yielded inactive Cdk4. In contrast, Src pretreatment of p27 did not affect p27-cyclin D1-Cdk4 complex formation. However, Src treatment led to tyrosine phosphorylation of p27 and catalytic activation of assembled cyclin D1-Cdk4-p27 complexes. Thus, while PKB-dependent p27 phosphorylation appears to increase cyclin D1-Cdk4-p27 assembly or stabilize these complexes in vitro, cyclin D1-Cdk4-p27 activation requires the tyrosine phosphorylation of p27. Constitutive activation of PKB and Abl or Src family kinases in cancers would drive p27 phosphorylation, increase cyclin D1-Cdk4 assembly and activation, and reduce the cyclin E-Cdk2 inhibitory function of p27. Combined therapy with both Src and PI3K/PKB inhibitors may reverse this process.

Association of Apparent Diffusion Coefficient with Disease Recurrence in Patients with Locally Advanced Cervical Cancer Treated with Radical Chemotherapy and Radiation Therapy

PURPOSE To investigate whether volumetrically derived apparent diffusion coefficient (ADC) from pretreatment diffusion-weighted (DW) magnetic resonance (MR) imaging is associated with disease recurrence in women with locally advanced cervical cancer treated with chemotherapy and radiation therapy. MATERIALS AND METHODS An ethics board-approved, retrospective study was conducted in 85 women with stage IB-IVA cervical cancer treated with chemo- and radiation therapy in 2009-2013. All patients underwent MR imaging for staging, including T2-weighted and DW MR imaging series, by using a 1.5- or 3.0-T imager. The mean, median, 75th, 90th, and 95th percentile ADCs (ADCmean, ADC50, ADC75, ADC90, and ADC95, respectively) of all voxels that comprised each tumor were extracted and normalized to the mean urine ADC (nADCmean, nADC50, nADC75, nADC90, and nADC95, respectively) to reduce variability. The primary outcome was disease-free survival (DFS). Uni- and multivariable Cox regression analyses were used to evaluate the association of ADC parameters and relevant clinical variables with DFS. RESULTS Of the 85 women included, 62 were free of disease at last follow-up. Median follow-up was 37 months (range, 5-68 months). Significant variables at univariable analysis included T2-weighted derived tumor diameter, para-aortic nodal involvement, advanced stage, ADC90 and ADC95, nADC75, nADC90, and nADC95. Normalized parameters were more highly associated (hazard ratio per 0.01 increase in normalized ADC, 0.91-0.94; P < .04). Because nADC75, nADC90, and nADC95 were highly correlated, only nADC95 (which had the lowest P value) was included in multivariable analysis. At multivariable analysis, absolute and normalized ADC95 remained associated with DFS (hazard ratio, 0.90-0.98; P < .05). CONCLUSION The volumetric ADC95 may be a useful imaging metric to predict treatment failure in patients with locally advanced cervical cancer treated with chemo- and radiation therapy.

Association between Metformin Use and Mortality after Cervical Cancer in Older Women with Diabetes

Background: To examine the association between metformin use and mortality in patients with diabetes and cervical cancer. Methods: Using Ontario health databases, a retrospective, population-based cohort study was conducted in women with diabetes ≥ age 66 years diagnosed with cervical cancer between 1997 and 2010. The association between metformin exposure and cervical cancer–specific mortality was examined using Fine–Gray regression models, with noncancer death as a competing risk and cumulative metformin use as a time-varying exposure. The association with overall mortality was examined using Cox regression models. Results: Among the 181 women with diabetes and cervical cancer, there were 129 deaths, including 61 cervical cancer–specific deaths. The median follow-up was 5.8 years (interquartile range 4.2–9.6 years) for surviving patients. Cumulative dose of metformin after cervical cancer diagnosis was independently associated with a decreased risk of cervical cancer–specific mortality and overall mortality in a dose-dependent fashion [HR 0.79; 95% confidence interval (CI), 0.63–0.98; and HR 0.95; 95% CI, 0.90–0.996 per each additional 365 g of metformin use, respectively]. There was no significant association between cumulative use of other antidiabetic drugs and cervical cancer–specific mortality. Conclusion: This study suggests an association between cumulative metformin use after cervical cancer diagnosis and lower cervical cancer–specific and overall mortality among older women with diabetes. Impact: Cumulative dose of metformin use after cervical cancer diagnosis among older women with diabetes may be associated with a significant decrease in mortality. This finding has important implications if validated prospectively, as metformin is inexpensive and can be easily combined with standard treatment for cervical cancer. Cancer Epidemiol Biomarkers Prev; 25(3); 507–12. ©2015 AACR.

In reply to Smith and Eifel.

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Brachytherapy in Gynecologic Cancers: Why Is It Underused?

Despite its established efficacy, brachytherapy is underused in the management of cervical and vaginal cancers in some parts of the world. Possible reasons for the underutilization of brachytherapy include the adoption of less invasive techniques, such as intensity-modulated radiotherapy; reimbursement policies favoring these techniques over brachytherapy; poor physician or patient access to brachytherapy; inadequate maintenance of brachytherapy skills among practicing radiation oncologists; transitioning to high-dose-rate (HDR) brachytherapy with increased time requirements; and insufficient training of radiation oncology residents.

Novel Imaging for Treatment Planning or Tumor Response

Anatomic imaging has long represented an integral part of modern radiotherapy, from planning, image-guidance to response evaluation. Functional imaging modalities now allow oncologists to supplement these anatomic images with functional maps, to elucidate the underlying biologic processes of cancer and allow delineation of both physical and biologic target volumes. Prescription of dose to this combination, commonly referred to as dose painting, represents an attractive avenue to further improve the therapeutic ratio of radiotherapy. This chapter focuses on the integration of novel imaging techniques and their role in delineating biologic radiotherapeutic targets, organized by well-known principles in radiobiology: tumor repopulation, reoxygenation, and repair. Focus is largely on clinically available imaging modalities, including positron emission tomography (PET) with various targeted radionuclides and functional magnetic resonance imaging (MRI). Other potential preclinical techniques are highlighted where relevant, particularly as they apply to promising translational concepts. Emphasis is placed on integration into treatment planning, adaptive treatment modification, and posttreatment response assessment.

The predictive value of nadir neutrophil count during treatment of cervical cancer: Interactions with tumor hypoxia and interstitial fluid pressure (IFP)

Background and purpose Hypoxia, high interstitial fluid pressure (IFP) and immune effects have individually been shown to modulate radiotherapy (RT) response in cervical cancer. The aim of this study was to investigate the interplay between hypoxia or IFP and circulating neutrophil levels, and their combined effect on survival following RT. Material and methods A total of 287 FIGO stage IB to IIIB cervical cancer patients treated with RT or RT and cisplatin (RTCT) were included. Tumor hypoxia and IFP were measured at baseline prior to treatment. Absolute neutrophil count (ANC) was measured at baseline and weekly during treatment. Median follow up was 7.1 years. Results High nadir ANC at the point of maximal myelosuppression was a stronger predictor of inferior survival than high baseline ANC after adjusting for clinical prognostic factors and treatment (RT vs. RTCT). The predictive effect of nadir ANC was most evident in patients with well-oxygenated tumors or tumors with high IFP at diagnosis. Conclusions This study provides new information about the combined influence of the tumor microenvironment and myeloid cells on the survival of cervical cancer patients treated with RT/RTCT to motivate the development of new treatments based on molecular targeting of immune–based radioresistance pathways.

Detection of extra-regional tumour recurrence with 18F-FDG-PET/CT in patients with recurrent gynaecological malignancies being considered for radical salvage surgery

AIM To compare the detection rate of extra-regional metastases in patients with recurrent gynaecological malignancies being considered for radical salvage surgery with combined 2-[18F]-fluoro-2-deoxy-d-glucose (18F-FDG) positron-emission tomography (PET)/computed tomography (CT) compared to conventional imaging. MATERIALS AND METHODS A retrospective review was performed of all patients in a provincial database with recurrent gynaecological malignancies being considered for pelvic exenteration that underwent restaging with CT/PET between March 2011 and October 2014. Findings on PET and conventional imaging (CT±pelvic MRI) were abstracted. Disease sites were classified according to the location (regional nodal metastases, extra-regional nodal metastases, peritoneum, or other distant sites) and diagnostic certainty (definitive or equivocal). The proportion of patients positive for extra-regional recurrence was calculated for PET and conventional imaging. In addition, the proportion of patients with indeterminate lesions only was calculated for each modality and the sites of indeterminate findings were tabulated. RESULTS There were 85 patients (median age, 50 years; range: 30-81 years) with carcinoma of cervix (n=51), endometrium (n=18), vagina (n=6), or vulva (n=10). Extra-regional recurrence was detected in 8/85 (9.4%) and 24/85 (28.2%) patients on conventional imaging and PET, respectively (p=0.0017). The greatest impact of PET compared to conventional imaging was in the detection of extra-regional nodal metastases. PET had significantly fewer examinations equivocal for extra-regional recurrence versus conventional imaging (6/85 [7.1%] and 38/85 [44.7%], respectively, p<0.001). CONCLUSION In patients with recurrent gynaecological malignancies being considered for radical salvage surgery, PET may identify significantly more extra-regional recurrence than conventional imaging. PET may also result in fewer equivocal lesions. The impact of these results on patient management and outcome should be confirmed in future prospective trials.

Comparison of dosimetric parameters derived from whole organ and wall contours for bladder and rectum in cervical cancer patients treated with intracavitary and interstitial brachytherapy

For volumes up to 2 cm3 of the bladder and possibly up to 5 cm3 of the rectum, doses computed from the whole organ were good estimates of the doses in the wall in cervix brachytherapy, and there were no significant differences between patients treated with or without interstitial needles.