Ryohei Sasaki

Mitochondrial respiration defects in cancer cells cause activation of Akt survival pathway through a redox-mediated mechanism

Cancer cells exhibit increased glycolysis for ATP production due, in part, to respiration injury (the Warburg effect). Because ATP generation through glycolysis is less efficient than through mitochondrial respiration, how cancer cells with this metabolic disadvantage can survive the competition with other cells and eventually develop drug resistance is a long-standing paradox. We report that mitochondrial respiration defects lead to activation of the Akt survival pathway through a novel mechanism mediated by NADH. Respiration-deficient cells (ρ-) harboring mitochondrial DNA deletion exhibit dependency on glycolysis, increased NADH, and activation of Akt, leading to drug resistance and survival advantage in hypoxia. Similarly, chemical inhibition of mitochondrial respiration and hypoxia also activates Akt. The increase in NADH caused by respiratory deficiency inactivates PTEN through a redox modification mechanism, leading to Akt activation. These findings provide a novel mechanistic insight into the Warburg effect and explain how metabolic alteration in cancer cells may gain a survival advantage and withstand therapeutic agents.

[18F]Fluorodeoxyglucose Uptake by Positron Emission Tomography Predicts Outcome of Non–Small-Cell Lung Cancer

PURPOSE To determine whether the standardized uptake value (SUV) of [(18)F]fluorodeoxyglucose uptake by positron emission tomography could be a prognostic factor for non-small-cell lung cancer (NSCLC). PATIENTS AND METHODS One hundred sixty-two patients with stage I to IIIb NSCLC were analyzed. Overall survival (OS), disease-free survival (DFS), distant metastasis-free survival (DMFS), and local-regional control (LRC) were calculated by the Kaplan-Meier method and evaluated with the log-rank test. The prognostic significance was assessed by univariate and multivariate analyses. RESULTS There were 93 patients treated with surgery and 69 patients treated with radiotherapy. A cutoff of 5 for the SUV for the primary tumor showed the best discriminative value. The SUV for the primary tumor was a significant predictor of OS (P = .02) in both groups. Low SUVs (</= 5.0) showed significantly better DFS rates than those with high SUVs (> 5.0; surgery group, P = .02; radiotherapy group, P = .0005). Low SUVs (</= 5.0) indicated a significantly better DFS than those with high SUVs (> 5.0; stage I or II, P = .02; stage IIIa or IIIb, P = .004). However, using the same cutoff point of 5, the SUV for regional lymph nodes was not a significant indicator for DFS (P = .19), LRC (P = .97), or DMFS (P = .17). The multivariate analysis showed that the SUV for the primary tumor was a significant prognostic factor for OS (P = .03) and DFS (P = .001). CONCLUSION The SUV of the primary tumor was the strongest prognostic factor among the patients treated by curative surgery or radiotherapy.

Novel role of p53 in maintaining mitochondrial genetic stability through interaction with DNA Pol γ

Mitochondrial DNA (mtDNA) mutations and deletions are frequently observed in cancer, and contribute to altered energy metabolism, increased reactive oxygen species (ROS), and attenuated apoptotic response to anticancer agents. The mechanisms by which cells maintain mitochondrial genomic integrity and the reason why cancer cells exhibit more frequent mtDNA mutations remain unclear. Here, we report that the tumor suppressor molecule p53 has a novel role in maintaining mitochondrial genetic stability through its ability to translocate to mitochondria and interact with mtDNA polymerase γ (pol γ) in response to mtDNA damage induced by exogenous and endogenous insults including ROS. The p53 protein physically interacts with mtDNA and pol γ, and enhances the DNA replication function of pol γ. Loss of p53 results in a significant increase in mtDNA vulnerability to damage, leading to increased frequency of in vivo mtDNA mutations, which are reversed by stable transfection of wild‐type p53. This study provides a mechanistic explanation for the accelerating genetic instability and increased ROS stress in cancer cells associated with loss of p53.

Reirradiation for locally recurrent lung cancer previously treated with radiation therapy

PURPOSE Local recurrence of lung cancer after previous external beam irradiation poses some problems for subsequent management. We retrospectively reviewed our series of patients with local recurrence of lung cancer to evaluate the efficacy and safety of reirradiation. PATIENTS AND METHODS Between 1979 and 2000, 34 patients with local recurrence of lung cancer were retreated with external radiation. There were 29 males and 5 females ranging in age from 38 to 85 years (median: 69 years). At the time of reirradiation, the clinical stage was I or II in 2 patients, IIIa in 5 patients, IIIb in 14 patients, and IV in 13 patients. Reirradiation was performed in 18 patients with the aim of achieving a cure or prolongation of survival (radical treatment), while 16 patients were treated for improvement of their symptoms (symptomatic treatment). RESULTS The median interval between the initial radiation therapy and reirradiation was 23 months, with a range of 5 to 87 months. The dose of initial irradiation delivered to the tumor ranged from 30 to 80 Gy (median: 60 Gy) in 1.5--2.0-Gy fractions per day. During reirradiation, it ranged from 10 to 70 Gy (median: 50 Gy) in 1.8--3.0-Gy fractions per day. The cumulative dose delivered to the tumor by treatments of both initial and second irradiation ranged from 56.5 to 150 Gy (median: 110 Gy). A response was observed in 14 out of 18 patients given radical treatment (complete response, 6; partial response, 8). Twelve of the 16 patients (75%) given symptomatic treatment also showed a symptomatic benefit. The overall survival rate after reirradiation was 43% at 1 year and 27% at 2 years, with a median survival time of 8 months. The median survival time after radical treatment was 15 months, with a range of 3 to 58 months, whereas that after symptomatic treatment was 3 months, with a range of 1 to 14 months. Six long-term survivors lived for more than 20 months. Reirradiation-induced toxicity included symptomatic radiation pneumonitis in 19 patients and symptomatic radiation esophagitis in 6 patients. These toxicities were not fatal, and radiation myelopathy was not caused by reirradiation. CONCLUSION Based on this study, external beam reirradiation can achieve satisfactory results for local recurrence of lung cancer provided that attention is paid to the possible hazards.

Compensatory glutamine metabolism promotes glioblastoma resistance to mTOR inhibitor treatment

The mechanistic target of rapamycin (mTOR) is hyperactivated in many types of cancer, rendering it a compelling drug target; however, the impact of mTOR inhibition on metabolic reprogramming in cancer is incompletely understood. Here, by integrating metabolic and functional studies in glioblastoma multiforme (GBM) cell lines, preclinical models, and clinical samples, we demonstrate that the compensatory upregulation of glutamine metabolism promotes resistance to mTOR kinase inhibitors. Metabolomic studies in GBM cells revealed that glutaminase (GLS) and glutamate levels are elevated following mTOR kinase inhibitor treatment. Moreover, these mTOR inhibitor-dependent metabolic alterations were confirmed in a GBM xenograft model. Expression of GLS following mTOR inhibitor treatment promoted GBM survival in an α-ketoglutarate-dependent (αKG-dependent) manner. Combined genetic and/or pharmacological inhibition of mTOR kinase and GLS resulted in massive synergistic tumor cell death and growth inhibition in tumor-bearing mice. These results highlight a critical role for compensatory glutamine metabolism in promoting mTOR inhibitor resistance and suggest that rational combination therapy has the potential to suppress resistance.

Mitochondrial DNA determines androgen dependence in prostate cancer cell lines

Prostate cancer progresses from an androgen-dependent to androgen-independent stage after androgen ablation therapy. Mitochondrial DNA plays a role in cell death and metastatic competence. Further, heteroplasmic large-deletion mitochondrial DNA is very common in prostate cancer. To investigate the role of mitochondrial DNA in androgen dependence of prostate cancers, we tested the changes of normal and deleted mitochondrial DNA in accordance with the progression of prostate cancer. We demonstrated that the androgen-independent cell line C4-2, established by inoculation of the androgen-dependent LNCaP cell line into castrated mice, has a greatly reduced amount of normal mitochondrial DNA and an accumulation of large-deletion DNA. Strikingly, the depletion of mitochondrial DNA from androgen-dependent LNCaP resulted in a loss of androgen dependence. Reconstitution of normal mitochondrial DNA to the mitochondrial DNA-depleted clone restored androgen dependence. These results indicate that mitochondrial DNA determines androgen dependence of prostate cancer cell lines. Further, mitochondrial DNA-deficient cells formed tumors in castrated athymic mice, whereas LNCaP did not. The accumulation of large deletion and depletion of mitochondrial DNA may thus play a role in the development of androgen independence, leading to progression of prostate cancers.

The efficacy of conventional radiation therapy in the management of pituitary adenoma

PURPOSE To evaluate the efficacy of conventional radiotherapy for reducing tumor size and endocrine hypersecretion of pituitary adenomas. METHODS AND MATERIALS We reviewed the records of 91 patients with pituitary adenoma, who were first treated between 1969 and 1994 and had been followed for more than 2 years (median, 8.2 years.) Of these patients, 86 had received postoperative radiotherapy, and 5 had received radiotherapy alone. The median total dose was 51 Gy. Clinical symptoms related to mass effects or endocrine hypersecretion were assessed. The efficacy of radiotherapy was evaluated before treatment and during the follow-up period (1-14 years; median, 3 years) by estimating tumor size on computed tomography or magnetic resonance imaging in 56 patients, as well as by endocrine testing in the 22 patients who had secreting adenomas. Local control rate, prognostic factors, and side effects were analyzed. RESULTS Mass-effect symptoms improved in 72% and 79% of patients who had such symptoms due to nonsecreting adenomas and secreting adenomas, respectively. Symptoms of endocrine hypersecretion abated in 67% of patients who had such symptoms. Excessive hormone levels normalized in 74% of patients who showed endocrine hypersecretions. The greatest size reduction was seen 3 years after the completion of radiotherapy (24% CR, 62% PR, 12% NC, and 3% PD in nonsecreting adenomas, and 32% CR, 36% PR, 27% NC, and 5% PD in secreting adenomas). Three patients with secreting adenomas (2 with prolactinoma and 1 with Cushings disease) showed a mismatch between reduction in tumor size and normalization of endocrine hypersecretion. The 10-year local control rates were 98%, 85%, 83%, and 67% for nonsecreting adenoma, growth-hormone-secreting adenoma, prolactinoma, and Cushings disease, respectively. Univariate analyses showed that disease type and radiation field size were significant prognostic factors. Brain necrosis occurred in 1 patient who received a 60-Gy dose of irradiation. CONCLUSION We conclude that conventional external radiotherapy with 50 Gy is safe and sufficient to control pituitary adenoma. Careful observation is required in the management of secreting adenomas because the effects on tumor size and endocrine hypersecretion may be mismatched in some secreting adenomas.

Clinical results and risk factors of proton and carbon ion therapy for hepatocellular carcinoma

The objective of this study was to evaluate the clinical outcome of proton and carbon ion therapy for hepatocellular carcinoma (HCC).

A phase I/II study of gemcitabine-concurrent proton radiotherapy for locally advanced pancreatic cancer without distant metastasis

PURPOSE We conducted the study to assess the feasibility and efficacy of gemcitabine-concurrent proton radiotherapy (GPT) for locally advanced pancreatic cancer (LAPC). MATERIALS AND METHODS Of all 50 patients who participated in the study, 5 patients with gastrointestinal (GI)-adjacent LAPC were enrolled in P-1 (50 Gy equivalent [GyE] in 25 fractions) and 5 patients with non-GI-adjacent LAPC in P-2 (70.2 GyE in 26 fractions), and 40 patients with LAPC regardless of GI-adjacency in P-3 (67.5 GyE in 25 fractions using the field-within-a-field technique). In every protocol, gemcitabine (800 mg/m(2)/week for 3 weeks) was administered concurrently. Every patient received adjuvant chemotherapy including gemcitabine after GPT within the tolerable limit. RESULTS The median follow-up period was 12.5 months. The scheduled GPT was feasible for all except 6 patients (12%) due to acute hematologic or GI toxicities. Grade 3 or greater late gastric ulcer and hemorrhage were seen in 5 patients (10%) in P-2 and P-3. The one-year freedom from local-progression, progression-free, and overall survival rates were 81.7%, 64.3%, and 76.8%, respectively. CONCLUSION GPT was feasible and showed high efficacy. Although the number of patients and the follow-up periods are insufficient, the clinical results seem very encouraging.

DNA polymerase γ inhibition by vitamin K3 induces mitochondria-mediated cytotoxicity in human cancer cells

Among the vitamin K (VK) compounds, VK3 exhibits distinct cytotoxic activity in cancer cells and is thought to affect redox cycling; however, the underlying mechanisms remain unclear. Here we demonstrate that VK3 selectively inhibits DNA polymerase (pol) γ, the key enzyme responsible for mitochondrial DNA replication and repair. VK3 at 30 µM inhibited pol γ by more than 80%, caused impairment of mitochondrial DNA replication and repair, and induced a significant increase in reactive oxygen species (ROS), leading to apoptosis. At a lower concentration (3 µM), VK3 did not cause a significant increase in ROS, but was able to effectively inhibit cell proliferation, which could be reversed by supplementing glycolytic substrates. The cytotoxic action of VK3 was independent of p53 tumor suppressor gene status. Interestingly, VK3 only inhibited pol γ but did not affect other pol including human pol α, pol β, pol δ, and pol ɛ. VK1 and VK2 exhibited no inhibitory effect on any of the pol tested. These data together suggest that the inhibition of pol γ by VK3 is relatively specific, and that this compound seems to exert its anticancer activity by two possible mechanisms in a concentration‐dependent manner: (1) induction of ROS‐mediated cell death at high concentrations; and (2) inhibition of cell proliferation at lower concentrations likely through the suppression of mitochondrial respiratory function. These findings may explain various cytotoxic actions induced by VK3, and may pave the way for the further use of VK3. (Cancer Sci 2008; 99: 1040–1048)