Targeting HSP90 as a Novel Therapy for Cancer: Mechanistic Insights and Translational Relevance.

Heat shock protein (HSP90), a highly conserved molecular chaperon, is indispensable for the maturation of newly synthesized poly-peptides and provides a shelter for the turnover of misfolded or denatured proteins. In cancers, the client proteins of HSP90 extend to the entire process of oncogenesis that are associated with all hallmarks of cancer. Accumulating evidence has demonstrated that the client proteins are guided for proteasomal degradation when their complexes with HSP90 are disrupted. Accordingly, HSP90 and its co-chaperones have emerged as viable targets for the development of cancer therapeutics. Consequently, a number of natural products and their analogs targeting HSP90 have been identified. They have shown a strong inhibitory effect on various cancer types through different mechanisms. The inhibitors act by directly binding to either HSP90 or its co-chaperones/client proteins. Several HSP90 inhibitors-such as geldanamycin and its derivatives, gamitrinib and shepherdin-are under clinical evaluation with promising results. Here, we review the subcellular localization of HSP90, its corresponding mechanism of action in the malignant phenotypes, and the recent progress on the development of HSP90 inhibitors. Hopefully, this comprehensive review will shed light on the translational potential of HSP90 inhibitors as novel cancer therapeutics.

Abdominal aortic aneurysm rupture presenting with focal weakness and altered mental status: a case report.

BACKGROUND: Abdominal aortic aneurysms (AAA) can present asymptomatically and may be found through routine screening or seen incidentally on imaging. Rupture due to weaking of the aortic wall is the main complication of an AAA and leads to approximately 200,000 deaths annually worldwide. Clinically, AAA rupture most frequently presents with abdominal and/or back pain, pulsatile abdominal mass, and hypotension. Here, we present an unusual presentation of embolic cerebrovascular accident associated with an AAA rupture. CASE PRESENTATION: A 58-year-old African American man transported to the emergency department via ambulance presents with altered mental status and unilateral extremity weakness. The initial presentation was concerning for acute cerebrovascular accident, acute kidney injury, severe sepsis, and urinary tract infection. Several hours after the initial presentation, the patient's abdomen began to appear distended and he became hypotensive. An abdominal CT was ordered which showed a large AAA rupture with a retroperitoneal bleed. The patient was transferred to a higher-level medical center for surgical repair. CONCLUSION: Abdominal aortic aneurysm rupture can rarely present due to an acute cerebrovascular accident with altered mental status and focal neurologic deficits.

Antitumor Activity of a Mitochondrial-Targeted HSP90 Inhibitor in Gliomas.

PURPOSE: To investigate the antitumor activity of a mitochondrial-localized HSP90 inhibitor, Gamitrinib, in multiple glioma models, and to elucidate the antitumor mechanisms of Gamitrinib in gliomas. EXPERIMENTAL DESIGN: A broad panel of primary and temozolomide (TMZ)-resistant human glioma cell lines were screened by cell viability assays, flow cytometry, and crystal violet assays to investigate the therapeutic efficacy of Gamitrinib. Seahorse assays were used to measure the mitochondrial respiration of glioma cells. Integrated analyses of RNA sequencing (RNAseq) and reverse phase protein array (RPPA) data were performed to reveal the potential antitumor mechanisms of Gamitrinib. Neurospheres, patient-derived organoids (PDO), cell line-derived xenografts (CDX), and patient-derived xenografts (PDX) models were generated to further evaluate the therapeutic efficacy of Gamitrinib. RESULTS: Gamitrinib inhibited cell proliferation and induced cell apoptosis and death in 17 primary glioma cell lines, 6 TMZ-resistant glioma cell lines, 4 neurospheres, and 3 PDOs. Importantly, Gamitrinib significantly delayed the tumor growth and improved survival of mice in both CDX and PDX models in which tumors were either subcutaneously or intracranially implanted. Integrated computational analyses of RNAseq and RPPA data revealed that Gamitrinib exhibited its antitumor activity via (i) suppressing mitochondrial biogenesis, OXPHOS, and cell-cycle progression and (ii) activating the energy-sensing AMP-activated kinase, DNA damage, and stress response. CONCLUSIONS: These preclinical findings established the therapeutic role of Gamitrinib in gliomas and revealed the inhibition of mitochondrial biogenesis and tumor bioenergetics as the primary antitumor mechanisms in gliomas.

A Modified Nucleoside 6-Thio-2'-Deoxyguanosine Exhibits Antitumor Activity in Gliomas.

PURPOSE: To investigate the therapeutic role of a novel telomere-directed inhibitor, 6-thio-2'-deoxyguanosine (THIO) in gliomas both in vitro and in vivo. EXPERIMENTAL DESIGN: A panel of human and mouse glioma cell lines was used to test therapeutic efficacy of THIO using cell viability assays, flow cytometric analyses, and immunofluorescence. Integrated analyses of RNA sequencing and reverse-phase protein array data revealed the potential antitumor mechanisms of THIO. Four patient-derived xenografts (PDX), two patient-derived organoids (PDO), and two xenografts of human glioma cell lines were used to further investigate the therapeutic efficacy of THIO. RESULTS: THIO was effective in the majority of human and mouse glioma cell lines with no obvious toxicity against normal astrocytes. THIO as a monotherapy demonstrated efficacy in three glioma cell lines that had acquired resistance to temozolomide. In addition, THIO showed efficacy in four human glioma cell lines grown as neurospheres by inducing apoptotic cell death. Mechanistically, THIO induced telomeric DNA damage not only in glioma cell lines but also in PDX tumor specimens. Integrated computational analyses of transcriptomic and proteomic data indicated that THIO significantly inhibited cell invasion, stem cell, and proliferation pathways while triggering DNA damage and apoptosis. Importantly, THIO significantly decreased tumor proliferation in two PDO models and reduced the tumor size of a glioblastoma xenograft and a PDX model. CONCLUSIONS: The current study established the therapeutic role of THIO in primary and recurrent gliomas and revealed the acute induction of telomeric DNA damage as a primary antitumor mechanism of THIO in gliomas.

Pathway signatures derived from on-treatment tumor specimens predict response to anti-PD1 blockade in metastatic melanoma.

Both genomic and transcriptomic signatures have been developed to predict responses of metastatic melanoma to immune checkpoint blockade (ICB) therapies; however, most of these signatures are derived from pre-treatment biopsy samples. Here, we build pathway-based super signatures in pre-treatment (PASS-PRE) and on-treatment (PASS-ON) tumor specimens based on transcriptomic data and clinical information from a large dataset of metastatic melanoma treated with anti-PD1-based therapies as the training set. Both PASS-PRE and PASS-ON signatures are validated in three independent datasets of metastatic melanoma as the validation set, achieving area under the curve (AUC) values of 0.45-0.69 and 0.85-0.89, respectively. We also combine all test samples and obtain AUCs of 0.65 and 0.88 for PASS-PRE and PASS-ON signatures, respectively. When compared with existing signatures, the PASS-ON signature demonstrates more robust and superior predictive performance across all four datasets. Overall, we provide a framework for building pathway-based signatures that is highly and accurately predictive of response to anti-PD1 therapies based on on-treatment tumor specimens. This work would provide a rationale for applying pathway-based signatures derived from on-treatment tumor samples to predict patients' therapeutic response to ICB therapies.

Targeting mTOR signaling overcomes acquired resistance to combined BRAF and MEK inhibition in BRAF-mutant melanoma.

Targeting MAPK pathway using a combination of BRAF and MEK inhibitors is an efficient strategy to treat melanoma harboring BRAF-mutation. The development of acquired resistance is inevitable due to the signaling pathway rewiring. Combining western blotting, immunohistochemistry, and reverse phase protein array (RPPA), we aim to understanding the role of the mTORC1 signaling pathway, a center node of intracellular signaling network, in mediating drug resistance of BRAF-mutant melanoma to the combination of BRAF inhibitor (BRAFi) and MEK inhibitor (MEKi) therapy. The mTORC1 signaling pathway is initially suppressed by BRAFi and MEKi combination in melanoma but rebounds overtime after tumors acquire resistance to the combination therapy (CR) as assayed in cultured cells and PDX models. In vitro experiments showed that a subset of CR melanoma cells was sensitive to mTORC1 inhibition. The mTOR inhibitors, rapamycin and NVP-BEZ235, induced cell cycle arrest and apoptosis in CR cell lines. As a proof-of-principle, we demonstrated that rapamycin and NVP-BEZ235 treatment reduced tumor growth in CR xenograft models. Mechanistically, AKT or ERK contributes to the activation of mTORC1 in CR cells, depending on PTEN status of these cells. Our study reveals that mTOR activation is essential for drug resistance of melanoma to MAPK inhibitors, and provides insight into the rewiring of the signaling networks in CR melanoma.

In perspective: An update on telomere targeting in cancer.

Engaging a telomere maintenance mechanism during DNA replication is essential for almost all advanced cancers. The conversion from normal and premalignant somatic cells to advanced malignant cells often results (85%-90%) from the reactivation of the functional ribonucleoprotein holoenzyme complex, referred to as telomerase. Modulation of the human telomerase reverse transcriptase (hTERT) appears to be rate limiting to produce functional telomerase and engage a telomere maintenance mechanism. The remaining 10% to 15% of cancers overcome progressively shortened telomeres by activating an alternative lengthening of telomeres (ALT) maintenance mechanism, through a DNA recombination pathway. Exploration into the specific mechanisms of telomere maintenance in cancer have led to the development of drugs such as Imetelstat (GRN163L), BIBR1532, 6-thio-dG, VE-822, and NVP-BEZ235 being investigated as therapeutic approaches for treating telomerase and ALT tumors. The successful use of 6-thio-dG (a nucleoside preferentially recognized by telomerase) that targets and uncaps telomeres in telomerase positive but not normal telomerase silent cells has recently shown impressive effects on multiple types of cancer. For example, 6-thio-dG overcomes therapy-resistant cancers in a fast-acting mechanism potentially providing an alternative or additional route of treatment for patients with cancer. In this perspective, we provide a synopsis of the current landscape of telomeres and telomerase processing in cancer development and how this new knowledge may improve outcomes for patients with cancer.

Induction of Telomere Dysfunction Prolongs Disease Control of Therapy-Resistant Melanoma.

Purpose: Telomerase promoter mutations are highly prevalent in human tumors including melanoma. A subset of patients with metastatic melanoma often fail multiple therapies, and there is an unmet and urgent need to prolong disease control for those patients.Experimental Design: Numerous preclinical therapy-resistant models of human and mouse melanoma were used to test the efficacy of a telomerase-directed nucleoside, 6-thio-2'-deoxyguanosine (6-thio-dG). Integrated transcriptomics and proteomics approaches were used to identify genes and proteins that were significantly downregulated by 6-thio-dG.Results: We demonstrated the superior efficacy of 6-thio-dG both in vitro and in vivo that results in telomere dysfunction, leading to apoptosis and cell death in various preclinical models of therapy-resistant melanoma cells. 6-thio-dG concomitantly induces telomere dysfunction and inhibits the expression level of AXL.Conclusions: In summary, this study shows that indirectly targeting aberrant telomerase in melanoma cells with 6-thio-dG is a viable therapeutic approach in prolonging disease control and overcoming therapy resistance. Clin Cancer Res; 24(19); 4771-84. ©2018 AACR See related commentary by Teh and Aplin, p. 4629.

Femur fractures in professional athletes: a case series.

OBJECTIVE: To discuss return to play after femur fractures in several professional athletes. BACKGROUND: Femur fractures are rare injuries and can be associated with significant morbidity and mortality. No reports exist, to our knowledge, on return to play after treatment of isolated femur fractures in professional athletes. Return to play is expected in patients with femur fractures, but recovery can take more than 1 year, with an expected decrease in performance. TREATMENT: Four professional athletes sustained isolated femur fractures during regular-season games. Two athletes played hockey, 1 played football, and 1 played baseball. Three players were treated with anterograde intramedullary nails, and 1 was treated with retrograde nailing. All players missed the remainder of the season. At an average of 9.5 months (range, 7-13 months) from the time of injury, all athletes were able to return to play. One player required the removal of painful hardware, which delayed his return to sport. Final radiographs revealed that all fractures were well healed. No athletes had subjective complaints or concerns that performance was affected by the injury at an average final follow-up of 25 months (range, 22-29 months). UNIQUENESS: As the size and speed of players increase, on-field trauma may result in significant injury. All players returned to previous levels of performance or exceeded previous statistical performance levels. CONCLUSIONS: In professional athletes, return to play from isolated femur fractures treated with either an anterograde or retrograde intramedullary nail is possible within 1 year. Return to the previous level of performance is possible, and it is important to develop management protocols, including rehabilitation guidelines, for such injuries. However, return to play may be delayed by subsequent procedures, including hardware removal.

Correlating MRI findings with disability in syndesmotic sprains of NFL players.

BACKGROUND: Syndesmotic sprains may be a significant source of missed playing time, especially in football players. Advanced imaging is frequently used to confirm the clinical diagnosis. Our purpose was to evaluate the prognostic ability of MRI in predicting time of disability. METHODS: Training room records from 1993 to 2007 for three National Football League teams were reviewed. Forty-three players were diagnosed with syndesmotic ankle injuries and underwent radiographs and magnetic resonance imaging. A blinded musculoskeletal radiologist interpreted all images. Players with fractures were excluded. RESULTS: Thirty-six professional football players were included in the final analysis. Twenty-three players had a positive squeeze test which was correlated with increased missed practices (p = 0.012) and increased missed games (p ≤ 0.01). The average number of games missed was 3.3 (range, 0 to 20) and the average number of practices missed was 16.7 (range, 0 to 114). Four players had isolated injury to the anterior tibio-fibular ligament (AITFL) (MRI Grade I). Five players had injury to the AITFL and interosseous ligament (MRI Grade II). Twenty-four players sustained injury to the AITFL, interosseous ligament, and posterior inferior tibio-fibular ligament (MRI Grade III). Three players had Grade III injuries with additional injury to the deltoid ligament (MRI Grade IV). Increasing grade of injury was positively correlated with increased number of missed games (p = 0.033) and missed practices (p = 0.002). CONCLUSION: MRI can be useful to help delineate the injury pattern and associated injuries, and may be useful in predicting time of disability using a grading system. Positive squeeze test can also be useful to determine prognosis.

Fluid consumption and sweating in National Football League and collegiate football players with different access to fluids during practice.

CONTEXT: Considerable controversy regarding fluid replacement during exercise currently exists. OBJECTIVE: To compare fluid turnover between National Football League (NFL) players who have constant fluid access and collegiate football players who replace fluids during water breaks in practices. DESIGN: Observational study. SETTING: Respective preseason training camps of 1 National Collegiate Athletic Association Division II (DII) football team and 1 NFL football team. Both morning and afternoon practices for DII players were 2.25 hours in length, and NFL players practiced for 2.25 hours in the morning and 1 hour in the afternoon. Environmental conditions did not differ. PATIENTS OR OTHER PARTICIPANTS: Eight NFL players (4 linemen, 4 backs) and 8 physically matched DII players (4 linemen, 4 backs) participated. INTERVENTION(S): All players drank fluids only from their predetermined individual containers. The NFL players could consume both water and sports drinks, and the DII players could only consume water. MAIN OUTCOME MEASURE(S): We measured fluid consumption, sweat rate, total sweat loss, and percentage of sweat loss replaced. Sweat rate was calculated as change in mass adjusted for fluids consumed and urine produced. RESULTS: Mean sweat rate was not different between NFL (2.1 +/- 0.25 L/h) and DII (1.8 +/- 0.15 L/h) players (F(1,12) = 2, P = .18) but was different between linemen (2.3 +/- 0.2 L/h) and backs (1.6 +/- 0.2 L/h) (t(14) = 3.14, P = .007). We found no differences between NFL and DII players in terms of percentage of weight loss (t(7) = -0.03, P = .98) or rate of fluid consumption (t(7) = -0.76, P = .47). Daily sweat loss was greater in DII (8.0 +/- 2.0 L) than in NFL (6.4 +/- 2.1 L) players (t(7) = -3, P = .02), and fluid consumed was also greater in DII (5.0 +/- 1.5 L) than in NFL (4.0 +/- 1.1 L) players (t(7) = -2.8, P = .026). We found a correlation between sweat loss and fluids consumed (r = 0.79, P < .001). CONCLUSIONS: During preseason practices, the DII players drinking water at water breaks replaced the same volume of fluid (66% of weight lost) as NFL players with constant access to both water and sports drinks.

Sweat rates and fluid turnover in professional football players: a comparison of National Football League linemen and backs.

CONTEXT: Many National Football League (NFL) teams practice 2 times per day over consecutive days in a hot and humid environment. Large body surface area (BSA) and use of protective equipment result in high sweat rates and total sweat loss in these football players. OBJECTIVE: To compare sweat rate, sweat loss, fluid consumption, and weight loss between NFL linemen and backs during preseason practices. DESIGN: Between-groups design. SETTING: Preseason training camp with wet bulb globe temperature between 19 degrees C and 25 degrees C. PATIENTS OR OTHER PARTICIPANTS: Eight linemen and 4 backs and receivers participated. MAIN OUTCOME MEASURE(S): Data were collected during both practice sessions on 2 separate days during the first week of August. Sweat rate was calculated as change in mass adjusted for all fluids consumed between prepractice and postpractice body mass measurements and the urine produced during practice divided by the length of the practice session. Gross daily sweat losses also were calculated. RESULTS: Height, mass, and BSA were higher in linemen than in backs. Sweat rate was also higher in linemen (2385 +/- 520 mL/h) than in backs (1410 +/- 660 mL/h, P < .001), as was the total volume of sweat lost during both practices in 1 day (6870 +/- 1034 mL/d versus 4110 +/- 2287 mL/d, P = .014). Compared with backs, linemen consumed more fluids during practice (2030 +/- 849 mL versus 1179 +/- 753 mL, P = .025) but produced less urine (53 +/- 73 mL versus 163 +/- 141 mL, P = .018). There was no difference in postpractice weight loss (linemen = -1.15 +/- 0.83%, backs = -1.06 +/- 0.76%). CONCLUSIONS: Linemen sweated at higher rates, lost larger volumes of sweat, consumed more fluids, and produced less urine during practice compared with the physically smaller backs, but they did not lose a greater percentage of body weight. Sodium losses could be considerable in NFL players during the preseason because of high daily sweat losses in backs and in linemen.

Core temperature and percentage of dehydration in professional football linemen and backs during preseason practices.

CONTEXT: Thermal responses of average-sized male subjects (mass of approximately 70 kg) may not accurately reflect the rate of heat storage in larger athletes with greater muscle mass. OBJECTIVE: To determine if core temperature (T(c)) is different in National Football League linemen and backs and if T(c) is related to percentage of dehydration or sweat rate. DESIGN: We measured T(c) and sweat rate in professional football players during preseason twice-daily practices. SETTING: Preseason training camp. PATIENTS OR OTHER PARTICIPANTS: Eight linemen (age = 26.6 +/- 2.1 years, height = 191.8 +/- 4.5 cm, mass = 134.8 +/- 10.7 kg, body surface area = 2.61 +/- 0.12 m2) and 6 backs (age = 27.0 +/- 4.2 years, height = 185.0 +/- 6.3 cm, mass = 95.6 +/- 11.1 kg, body surface area = 2.19 +/- 0.16 m2). MAIN OUTCOME MEASURE(S): We measured T(c) using ingestible sensors. Resting T(c) was recorded in the mornings of data collection with players dressed in shorts and then every 15 minutes during 2-hour practices in full pads or shells. Mass was recorded before and after practices for determining the percentage of dehydration. In 8 of the 14 subjects (4 linemen, 4 backs), sweat rate was calculated using the change in mass adjusted for fluid intake and urine production. RESULTS: Height, mass, and body surface area were greater in linemen than in backs. We noted a linear trend over time for T(c) in both groups. Maximal T(c) was higher in linemen (38.65 +/- 0.48 degrees C) than in backs (38.44 +/- 0.32 degrees C), but linemen were less dehydrated than backs (-0.94 +/- 0.6% versus -1.3 +/- 0.7%). Sweat rate was 2.11 +/- 0.77 L/h and correlated significantly with body surface area (r = 0.77, P < .05). Maximal T(c) was not correlated with either percentage of dehydration or sweat rate. CONCLUSIONS: Maximal T(c) was not associated with percentage of dehydration or sweat rate. Linemen were less dehydrated but demonstrated higher T(c) than backs during practice. Maximal T(c) was generally achieved during live scrimmaging.