MiR-23b/-27b Cluster Suppresses the Metastatic Phenotype of Aggressive Prostate Cancer Cells

MicroRNAs (miRs) are small, endogenous, non-coding RNAs that regulate gene expressing by inhibiting the stability and/or translation of complementary mRNA targets. Expression of miR-23b/-27b, which are encoded in tandem as a cluster on chromosome 9, are specifically downregulated in metastatic prostate cancer tissue samples compared to benign prostate. MiR-23b/-27b levels are also lower in aggressive prostate cancer cell lines compared to more indolent cells. We show miR-23b/-27b suppress migration and invasion of two aggressive human prostate cancer cell lines. Furthermore, inhibition of miR-23b/-27b increases the migration and invasion potential of a less aggressive prostate cancer cell line. Interestingly, we found that the miR-23b/-27b cluster does not affect prostate cancer cell proliferation suggesting miR-23b/-27b are metastatic suppressors. We provide evidence that miR-23b/-27b suppress additional key tumorigenic processes. To understand the mechanism of miR-23b/-27b mediated suppression of metastatic cell processes, we examined the effects of miR-23b/-27b on Rac1, a Rho GTPase essential for the metastatic phenotype of prostate cancer cells. We show ectopic expression of miR-23b/-27b decrease Rac1 activity of aggressive prostate cancer cell lines while inhibition of these miRs increase Rac1 activity in a less aggressive prostate cancer cell line. In addition, we show that expression of miR-23b/-27b increase the cell adhesion protein E-cadherin, in two independent aggressive prostate cancer cell lines. Conversely inhibition of miR-23b/-27b in a less aggressive prostate cancer cell line decreases E-cadherin levels. These data establish an important role for miR-23b/-27b mediated-suppression of invasion and migration of aggressive prostate cancer cells. Since miRs have great potential use as biomarkers and drug targets or mimics, elucidating the role and mechanism of miR-23b/-27b in prostate cancer is critical in the development of novel and effective therapies and prognostic indicators.</p

Massive hydraulic fracturing well, Federal No. 498-4-1, Rio Blanco County, Colorado. Final report

This project is an MHF of a previously untreated Mesaverde interval in a well in northwest Colorado. The rocks involved may have been deposited during a marine invasion of long-continued, swamp environments. If so, they would have possessed superior primary reservoir properties. The logging program, identical to those used in the nearby Rio Blanco Nuclear and MHF Project wells, supplied contradictory information. The frac could furnish better understanding of the log suite, better parameters for pre-frac judgments of productive potential and further proof of the commercial capabilities of the formation. The frac job was performed as designed. A total of 775,000 lbs of sand in a total of 276,000 gallons of gelled water (YF4PSD) were injected. The injection pressures ranged from 2,000 to 1,300 psig at rates from 37 to 10 BPM. During the post-frac clean up, 30% of the frac fluid flowed back in 36 hours. Following eight days of swabbing and a total fluid recovery of 46%, the well began continuous flow, which within an additional three days brought total fluid recovery to 70%. Then gas flow increased from gas-cut water to 800 MSCF/D and declined to about 200 MSCF/D within 22 days. After over four months of production and cumulative frac fluid recovery of 82%, the gas rate appears to stabilize at around 130 MSCF/D with about 7 BF/D which contain over 50% oil. The post-frac to pre-frac production ratio is 2:1. It is concluded that revision of interpretive methods and/or logging programs is required if consistency in selecting zones most productive of gas and most receptive to stimulation is to be achieved. Injecting large volumes into short vertical intervals appears presently economically unattractive in this area. Properly designed moderate volume multiple fracs may achieve commercial deliverability

Minocycline-Induced Agranulocytosis Presenting as Ecthyma Gangrenosum

A 51-year-old female with a history of rheumatoid arthritis was admitted for progressive fevers, chills and malaise. Five weeks prior, she started minocycline for an RA exacerbation. Two weeks after starting minocycline she developed an abscess on her right ankle that was treated at an urgent care facility with ceftriaxone and trimethoprim-sulfamethoxazole. She had minimal improvement so was switched to clindamycin. She developed additional abscesses on her right ankle and right axilla and spiking fevers so she was treated with incision and drainage under general anesthesia. Routine blood work obtained prior to surgery revealed severe neutropenia (0.74 103/ul) and the patient was urgently referred to the emergency department.  Skin biopsy was obtained on admission and revealed ulceration, necrosis, acute and chronic inflammation, vasculitis with vascular thrombosis and rod-shaped bacteria in blood vessel walls and lumina consistent with ecthyma gangrenosum. The following day tissue and blood cultures confirmed the growth of Pseudomonas aureginosa. Bone-marrow biopsy showed decreased granulopoiesis and hematopoiesis, and a diagnosis of minocycline-induced agranulocytosis presenting as ecthyma gangrenosum was made.  The patient had dramatic improvement with appropriate antibiotic therapy, discontinuation of minocycline and initiation of filgrastrim. She has remained healthy without recurrence for 17 months.   

Anonymity of Willed Body Donors: Survey Results from First Year Medical Students

There is current debate about the value of anonymity in willed body donors for first year medical students. Some claim that allowing the students to know more about the donors can enhance their empathy and can provide early experience in the doctor – patient relationship. Others claim that knowing too much about the donors may make the dissection experience more traumatic and can take away from the student's anatomical learning experience. To gain a student's perspective on this issue, first year medical students in two different curricular tracks at the University of Miami Miller School of Medicine were surveyed both before and after they completed their first year gross anatomy course (IRB 20160480). One curricular track provides 150 students with a short, eight week, high intensity anatomy course while the other track provides 50 students with a longer, 6 month, low intensity course. The online survey asked the students to rate how much personal information about the donors they would like to receive. The survey response rate was 76% and 63% for the pre‐ and post‐ short anatomy course with response rates of 58% and 56% for the pre‐ and post‐ long anatomy course. There was no difference in the results between the two curricular tracks and no difference in the results before or after taking the anatomy course. The results found that the students would prefer to have information about the donors that includes their medical history, social history and socioeconomic status (3.7 mean Likert score), but are less interested in having complete information about the donors (name, religion, political affiliation) (2.4 mean Likert score). Comments by the students suggested that knowing too much information could have a negative impact on their ability to perform invasive dissections and could have negative emotional effects. The students felt, however, that knowing the donor's medical and social history would allow them to appreciate and synthesize the anatomical information to a greater degree. These results indicate that students would like more pertinent medical information about their donors, but that they have concerns about knowing the complete information about their willed body donors. This is from the Experimental Biology 2018 Meeting. There is no full text article associated with this published in The FASEB Journal

The microRNA -23b/-27b cluster suppresses the metastatic phenotype of castration-resistant prostate cancer cells.

MicroRNAs (miRs) are small, endogenous, non-coding RNAs that regulate the stability and/or translation of complementary mRNA targets. MiRs have emerged not only as critical modulators of normal physiologic processes, but their deregulation may significantly impact prostate and other cancers. The expression of miR-23b and miR-27b, which are encoded by the same miR cluster (miR-23b/-27b), are downregulated in metastatic, castration-resistant tumors compared to primary prostate cancer and benign tissue; however, their possible role in prostate cancer progression is unknown. We found that ectopic expression of miR-23b/-27b in two independent castration-resistant prostate cancer cell lines resulted in suppression of invasion and migration, as well as reduced survival in soft agar (a measure of anoikis). However, there was no effect of miR-23b/-27b on cell proliferation suggesting that these miRs function as metastasis (but not growth) suppressors in prostate cancer. Conversely, inhibition of miR-23b/-27b in the less aggressive androgen-dependent LNCaP prostate cancer cell line resulted in enhanced invasion and migration also without affecting proliferation. Mechanistically, we found that introduction of miR-23b/-27b in metastatic, castration-resistant prostate cancer cell lines resulted in a significant attenuation of Rac1 activity without affecting total Rac1 levels and caused increased levels of the tumor suppressor E-cadherin. Inhibition of these miRs had the opposite effect in androgen-dependent LNCaP cells. These results suggest that miR-23b/-27b are metastasis suppressors that might serve as novel biomarkers and therapeutic agents for castration-resistant disease

Abstract 1467: The miR-23b/-27b cluster decreases metastasis of aggressive prostate cancer

Abstract Metastasis is responsible for the vast majority of prostate cancer deaths. As such, it is essential to understand the mechanisms driving prostate cancer to this lethal stage. Deregulation of microRNAs is increasingly implicated in the progression and metastasis of prostate and other cancers. MiR-23b and miR-27b, two members of the same miR cluster (miR-23b/-27b), are down-regulated in human metastatic, castration resistant prostate cancer (CRPC) as compared to primary tumors and benign tissue. However, the role of the miR-23b/-27b cluster is not fully understood, particularly in metastatic disease. Interpretation of existing data is complicated by the analysis of the individual effects of miR-23b and miR-27b even though these miRs are co-transcribed and coordinately expressed. The primary goal of our study is to determine the effects of miR-23b/-27b on metastatic processes in vitro and in vivo. Ectopic expression of miR-23b/-27b in two aggressive prostate cancer cell lines decreases migration, invasion and anchorage-independent growth. Conversely, inhibition of miR-23b/-27b using specific antagomirs in relatively indolent prostate cancer cells promotes increased invasion and migration. E-cadherin protein and mRNA levels were inversely related to miR-23b/-27b levels. In contrast, manipulation of miR-23b/-27b levels had no effect on prostate cancer cell proliferation in any of the tested cell lines. These findings suggest that miR-23b/-27b is specifically linked to metastasis suppression. The Rho GTPase, Rac1, is hyperactive in CRPC cell lines and patient samples. Since Rac1 promotes invasion and migration, we investigated the effects of miR-23b/-27b expression and inhibition on Rac1 activity. Ectopic expression of miR-23b/-27b in aggressive prostate cancer cell lines significantly attenuates active Rac1 while having no effect on total Rac1 levels. Conversely, inhibition of miR-23b/-27b in less aggressive prostate cancer cells increased Rac1 activity but not Rac1 levels. We further examined the effects of miR-23b/-27b on prostate cancer metastasis in vivo. Metastatic prostate cancer cells expressing luciferase and miR-23b/-27b or a scrambled control, were injected into the ventral prostates of nude mice. Orthotopic tumor formation and metastases were assessed by bioluminescence imaging. Metastatic tumor burden was greatly decreased in the tumors derived from miR-23b/-27b expressing cells. Taken together, these data demonstrate that expression of miR-23b/-27b exerts metastasis-suppressing effects in vitro and in vivo. The miR-23b/-27b cluster may be a useful biomarker of poor prognosis in addition to having therapeutic potential in advanced, metastatic prostate cancer. Citation Format: Meghan A. Rice, Reema Ishteiwy, Thirupandiyur Udayakumar, Derek Dykxhoorn, Kerry L. Burnstein. The miR-23b/-27b cluster decreases metastasis of aggressive prostate cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 1467. doi:10.1158/1538-7445.AM2014-1467</jats:p

MiR-23b/

<p>-<b>27b significantly increases E-cadherin expression in castration –resistant cell lines.</b> ALVA31 or PC3-ML cells expressing miR-23b/-27b or transduced with scrambled control and LNCaP cells transfected with antagomiR-23b/-27b or control antagomiR were subjected to western blotting for E-cadherin and actin. Representative blots are shown of a total of 2–6 experiments.</p

MiR-23b/

<p>-<b>27b significantly decreases Rac1 activity but not total Rac1 levels in castration-resistant prostate cancer cell lines.</b> Rac1 activity assays were performed on ALVA31 (A) and PC3-ML (B) cells expressing miR-23b/-27b or transduced with scrambled control. GTP-bound Rac1 was separated from GDP-Rac1 using a pull-down assay as described in Materials and Methods. Complexes containing GTP-Rac1 (active Rac1) were collected, denatured and resolved by SDS-PAGE followed by Western blotting with anti-Rac1 antibodies. Total Rac1 (GDP-and GTP-bound) represents 5% of the original cell lysate. Actin was used as a loading control. Quantification of three independent experiments is shown with error bars representing SEM (*P<0.05), (**P<0.01).</p