Perfusion and diffusion MRI signatures in histologic and genetic subtypes of WHO grade II–III diffuse gliomas

Non-vestibular cranial nerve schwannomas (NVCNS) are rare lesions, representing <10?% of cranial nerve schwannomas. The optimal treatment for NVCNS is often derived from vestibular schwannomas experience. Surgical resection has been referred to as the first line treatment for those benign tumors, but significant complication rates are reported. Stereotactic radiosurgery (SRS) has arisen as a mainstay of treatment for many benign tumors, including schwanommas. We retrospectively reviewed the outcomes of NVCNS treated by SRS to characterize tumor control, symptom relief, toxicity, and the role of hypo-fractionation of SRS dose. Eighty-eight (88) patients, with ninety-five (95) NVCNS were treated with either single or multi-session SRS from 2001 to 2014. Local control was achieved in 94?% of patients treated (median follow-up of 33 months, range 1–155). Complications were seen in 7.4?% of cases treated with SRS. At 1-year, 57?% of patients had improvement or resolution of their symptoms, while 35?% were stable and 8?% had worsening or increased symptoms. While 42?% received only one session, results on local control were similar for one or multiple sessions (p?=?0.424). SRS for NVCNS is a treatment modality that provides excellent local control with minimal complication risk compared to traditional neurosurgical techniques. Tumor control obtained with a multi-session treatment was not significantly different from single session treatment. Safety profile was also comparable for uni or multi-session treatments. We concluded that, as seen in VS treated with CK SRS, radiosurgery treatment can be safely delivered in cases of NVCNS.     

High expression of the stem cell marker nestin is an adverse prognostic factor in WHO grade II–III astrocytomas and oligoastrocytomas

Infiltrating astrocytomas and oligoastrocytomas of low to anaplastic grade (WHO grades II and III), in spite of being associated with a wide range of clinical outcomes, can be difficult to subclassify and grade by the current histopathologic criteria. Unlike oligodendrogliomas and anaplastic oligodendrogliomas that can be identified by the 1p/19q codeletion and the more malignant glioblastomas (WHO grade IV astrocytomas) that can be diagnosed solely based on objective features on routine hematoxylin and eosin sections, no such objective criteria exist for the subclassification of grade II–III astrocytomas and oligoastrocytomas (A+OA II–III). In this study, we evaluated the prognostic and predictive value of the stem cell marker nestin in adult A+OA II–III (n = 50) using immunohistochemistry and computer-assisted analysis on tissue microarrays. In addition, the correlation between nestin mRNA level and total survival was analyzed in the NCI Rembrandt database. The results showed that high nestin expression is a strong adverse prognostic factor for total survival (p = 0.0004). The strength of the correlation was comparable to but independent of the isocitrate dehydrogenase 1/2 (IDH 1/2) mutation status. Histopathological grading and subclassification did not correlate significantly with outcome, although the interpretation of this finding is limited by the fact that grade III tumors were treated more aggressively than grade II tumors. These results suggest that nestin level and IDH 1/2 mutation status are strong prognostic features in A+OA II–III and possibly more helpful for treatment planning than routine histopathological variables such as oligodendroglial component (astrocytoma vs. oligoastrocytoma) and WHO grade (grade II vs. III).     

Long-term follow-up results of postoperative radiation therapy for Cushing’s disease

Objectives Radiotherapy is currently used in patients with residual or recurrent pituitary adenomas after surgery. However, there is little information of long-term outcome of patients with Cushing’s disease following radiotherapy. We assessed the long-term efficacy and toxicity of conventional radiotherapy in the control of Cushing’s disease after unsuccessful transsphenoidal surgery. Patients and Methods Forty patients with Cushing’s disease were treated with conventional external beam radiotherapy at our Institution between 1988 and 2002. The median age was 38. All patients received radiotherapy following unsuccessful surgery or at tumour recurrence to a dose of 45–50 Gy in 25–28 fractions. The persistence of active disease after surgery was diagnosed by the increased high plasma cortisol levels, high 24 h urinary cortisol levels and absence of cortisol suppression after administration of dexamethasone. Results The 5 and 10 year local tumour control was 93% and the 5 and 10 year survival was 97 and 95%. Normalization of plasma cortisol was seen in 28% of patients at 1 year, 73% at 3 years, 78% at 5 years and 84% at 10 years. The average timing to remission was 24 months. The most common side effect was hypopituitarism that increased progressively during the follow-up, being present in 62% and in 76% of patients at 5 and 10 years after RT. There were no other serious complications as radiation induced optic neuropathy or second tumours. Conclusion Radiotherapy is effective in the long-term tumour- and hormone hypersecretion control of ACTH-secreting pituitary adenomas, however with a high prevalence of hypopituitarism. At the moment, it remains an important treatment option after failure of surgery.     

Necrosis is a consistent factor to recurrence of meningiomas: should it be a stand-alone grading criterion for grade II meningioma?

The purpose of this study was to evaluate spontaneous necrosis as a possible isolated factor for progression and recurrence in grade I meningiomas classified according to the current World Health Organization (WHO) classification. Meningiomas are the most frequently reported primary intracranial tumours, accounting for more than 35%. The 2016 WHO classification of central nervous system tumors stratifies meningiomas in grades I (benign), II (atypical), and III (malignant), according to histopathological aspects and the risk of progression or recurrence. Among 110 patients with intracranial meningiomas, 70 were WHO grade I meningiomas with no findings of atypia (G1WON), 15 were WHO grade I with necrosis (G1WN), 21 were WHO grade II (G2), and 4 were WHO grade III (G3). The mean follow-up was 5.9?±?0.2 years. High performance scale (KPS?≥?80) was different (p?<?0.001) between WHO grade I meningiomas without (81.4%) and with (60%) necrosis. The 5-year mortality rate was 1.4, 6.7 and 5.9% for G1WON, G1WN and G2, respectively, with significant difference (p?=?0.011) related to the presence of necrosis. The risk of recurrence was 3.7 times higher in G1WN than in G1WON (p?=?0.017), and 4.2 times in G2 (p?=?0.010). Progression-free survival (PFS) was clearly higher in patients with G1WON compared to G1WN and G2 (p?=?0.002 and p?<?0.001, respectively). There was no significant difference in PFS between G1WN and G2 (p?=?0.692). Retreatment was also superior in meningioma with necrosis. Our findings provide clear statistical data to consider that patients with benign meningiomas and histologic findings of spontaneous necrosis are at increased risk of progression and recurrence compared to those with benign lesion without atypical features. Statistical analysis curves also suggest that these lesions behave more similarly to those currently classified as WHO grade II meningioma.     

Phase II study of preoperative radiotherapy and concomitant weekly intravenous oxaliplatin combined with oral capecitabine for stages II–III rectal cancer

Introduction

A prospective phase II study was conducted to assess the clinical activity and tolerability of oxaliplatin, capecitabine, and radiotherapy (RT) for neoadjuvant therapy of stages II?CIII rectal cancer.

Materials and methods

Patients with histologically confirmed stages II?CIII (T3?CT4 and/or N+) resectable rectal adenocarcinoma were eligible. Capecitabine was administered at 825?mg/m² twice daily for 5?days/week and oxaliplatin at 50?mg/m² on day 1 weekly for 5?weeks starting the first day of RT (before RT). RT consisted of a total dose of 45?Gy delivered in 25 fractions of 1.8?Gy, 5?days per week, for 5?weeks.

Results

A total of 46 patients were included (35 male, 10 female, median age 62?years). TNM Stage was T3 in 43 patients and T4 in 2. Twenty-eight patients had suspected nodal involvement. The intended chemoradiation treatment was completed in 94?% patients. Grade 3/4 toxicity included lymphocytopenia (6 patients), diarrhea (4 patients), emesis (2 patients), asthenia (3 patients), anorexia (1 patient), and hepatic toxicity (1 patient). Grade 1 neurotoxicity occurred in 18 patients, Grade 2 neurotoxicity in 3, and Grade 1 palmoplantar erythrodysesthesia in 2. Forty-two patients underwent surgery (complete resection 95?%, sphincter-saving operation 55?%). The overall pathologic response rate was 83?%, with a pathologic complete response (pCR) rate of 11.9?% (95?% CI 4.0?C25.6).

Conclusions

The pCR rate observed with oxaliplatin plus capecitabine and RT did not reach the pre-specified criteria of efficacy in this trial, which is in line with recent results of randomized phase III trials.     

Response,toxicity, failure patterns,and survival in five radiation therapy oncology group (RTOG) trials of sequential and/or concurrent chemotherapy and radiotherapy for locally advanced non–small-cell carcinoma of the lung

Purpose: The purpose of this study was to assess response, toxicity, failure patterns, and survival differences in three chemotherapy (ChT)/radiation therapy (RT) sequencing strategies for locally advanced non-small cell lung cancer (NSCLC).Methods and Materials: Five completed Radiation Therapy Oncology Group (RTOG) trials for Stage II-IIIA/B inoperable NSCLC patients employed one of the three following strategy groupings: 1) sequential ChT followed by standard RT (60 Gy in 6 weeks); 2) combined sequential and concurrent ChT and standard RT (60 Gy in 6 weeks); or 3) concurrent ChT and hyperfractionated RT (69.6 Gy in 6 weeks). All five trials required KPS ≥ 70; two trials (314 patients) required <5% weight loss and three trials (147 patients) had no minimum weight loss requirement. In all five trials the ChT used cisplatin with either vinblastine or oral etoposide. Combining data for the five trials yielded an evaluable group of 461 patients. The three methods of sequencing ChT and RT were evaluated for differences in response, acute and late toxicity, patterns of failure, and survival. Acute toxicity was defined as that occurring within 90 days from the start of RT. Late toxicity was defined as that occurring after 90 days from the start of RT. Acute or late toxicity ≥ grade 3 was defined as severe. Site of first failure was recorded by date. In-field failure excluded distant metastasis as a failure and included only tissue in the RT treatment field. Overall progression-free survival (PFS) was defined as survival without evidence of intra- or extrathoracic tumor or death from any cause.Results: Group 1 had a lower overall response rate (63%) compared to either Group 2 (77%) or Group 3 (79%), p = 0.03 and 0.003, respectively. Overall grade 4/5 acute toxicities were nearly equal between groups. The severe nonhematologic acute toxicities were significantly different by strategy group (p < 0.0001). Group 1 and 2 were not statistically different. Group 3 had significantly more patients with severe acute nonhematologic toxicity (55%) than either Group 1 (27%) or 2 (34%) with p < 0.0001 and p = 0.0005, respectively. This was due to a severe acute esophagitis rate of 34% for Group 3 versus 1.3% for Group 1 and 6% for Group 2 (p < 0.0001 for both comparisons). Overall grade 4/5 late toxicities did not differ by group. Severe late nonhematologic toxicities were different by group (p = 0.0098). Group 1 patients had significantly fewer severe late nonhematologic toxicities (14%) compared to patients in Groups 2 (26%) or 3 (28%) (p = 0.046 and 0.038, respectively). Severe late lung toxicity was 10% for Group 1 compared to 21% and 20% for Groups 2 and 3, respectively. Severe late lung toxicities differed by group (p = 0.033), but not severe late esophagitis (p = 0.077). There were no differences between the three strategy groups for patterns of first failure. The in-field failures were higher in Group 2 (71%) compared to Groups 1 (56%) and 3 (55%), p = 0.0478. Pairwise comparisons yielded p-values of 0.068 and 0.015 for Group 2 versus 1 and Group 2 versus 3, respectively. Three-year PFS was better in Group 2 (15%) and 3 (15%) compared to Group 1 (7%), but not statistically significant (p = 0.454). Similarly, in-field PFS was better in Group 2 (17%) and 3 (20%) than Group 1 (9%), but not significant (p = 0.167). There were improvements in 3-year survival for Group 2 (17%) and Group 3 (25%) compared to Group 1 (15%), but the differences were not statistically significant (p = 0.47). The same results were present for patients with less than 5% weight loss and patients with stage IIIA tumors.Conclusion: Thus, concurrent ChT and hyperfractionated RT had a higher incidence of severe acute esophageal toxicity. Severe late lung toxicity with concurrent ChT/hyperfractionated RT, as well as with induction ChT followed by concurrent ChT/standard RT, may be greater compared to sequential ChT/RT. Also, concurrent ChT/hyperfractionated RT, and induction ChT followed by concurrent ChT/standard RT, appear to have a benefit in terms of increased tumor response compared to sequential ChT/RT, but not in failure pattern. The latter may reflect the presence of pretherapy micrometastases that are delayed from progression by the combined therapy, but not prevented.     

Does the extent of surgery have an impact on the survival of patients who receive postoperative radiation therapy for supratentorial low‐grade gliomas?

We evaluate the impact of extent of surgery (EOS) on survival of patients with supratentorial nonpilocytic low‐grade gliomas (LGG) treated with postoperative radiation therapy (PORT). Sixty‐five patients with pathologically confirmed supratentorial nonpilocytic LGG (36 astrocytomas and 29 oligodendrogliomas) were treated with PORT after different extents of surgery: 12 gross total resections (GTR), 27 minimal or subtotal resections (MR/SR), and 26 biopsies (B). EOS was confirmed with postoperative imaging. The median radiation dose delivered was 5,940 cGy (range, 4,950–6,620 cGy). One of 12 patients (8%) in the GTR group and 12 of 53 patients (23%) in the less than GTR group demonstrated contrast enhancement. The median follow‐up was 61 months (range 5–194 month). The 10‐year overall survival (OS) was 82.5% and 32% for the GTR and the less than GTR groups, respectively (P = 0.0008). The corresponding 10‐year disease‐specific survival (DSS) was 90% and 41.4%%, respectively (P = 0.001). Multivariate analysis showed that only contrast enhancement and EOS were predictors for OS and DSS. Our data suggest that EOS correlates with OS and DSS in patients who have PORT. GTR should be the goal if technically achievable without causing significant morbidity, and its combination with PORT is compatible with long‐term survival. © 2002 Wiley‐Liss, Inc.     

Antiepileptic and psychiatric medication in a nationwide cohort of patients with glioma WHO grade II–IV

Journal of Neuro-Oncology - Glioma is the most common intracranial primary brain tumor. Patients with glioma often suffer from epilepsy, anxiety and depression. Aims of this study were to identify...     

Differential expression of miR200a-3p and miR21 in grade II–III and grade IV gliomas: Evidence that miR200a-3p is regulated by O6-methylguanine methyltransferase and promotes temozolomide responsiveness

Glioblastoma multiforme (GBM) is the most common primary brain tumor and is among the deadliest of human cancers. Dysregulation of microRNAs (miRNAs) expression is an important step in tumor progression as miRNAs can act as tumor suppressors or oncogenes and may affect cell sensitivity to chemotherapy. Whereas the oncogenic miR21 has been shown to be overexpressed in gliomas, the expression and function of the tumor-supressor miR200a in GBMs remains unknown. In this study, we show that miR21 is upregulated in grade IV (GBMs) vs. grade II–III (LGs) gliomas, confirming that miR21 expression level is correlated with tumor grade, and that it may be considered as a marker of tumor progression. Conversely, miR200a is demonstrated for the first time to be downregulated in GBMs compared with LGs, and overexpression of miR200a in GBM cells is shown to promote TMZ-sensitivity. Interestingly, miR200a but not miR21 expression level is significantly higher in TMZ-responsive vs. -unresponsive tumoral glial cells in primary culture. Furthermore, miR200a appears negatively correlated with the expression of the DNA repair enzyme O⁶-methylguanine methyltransferase (MGMT), and the inhibition of MGMT activity results in an increase of miR200a expression in GBM cells. Taken together, these data strongly suggest that miR200a is likely to act as a crucial antitumoral factor regarding glioma progression. Interplay between miR200a and MGMT should be considered as potential mechanism involved in therapeutic response.     

A phase I/II study to evaluate radiation therapy and hyperthermia for deep-seated tumours: A report of RTOG 89–08

The purpose of this paper is to evaluate the safety and efficacy of deep hyperthermia in conjunction with radiation therapy. This study employed ‘second generation’ electromagnetic devices which were felt to be better able to confine heating and spare normal tissue than the devices evaluated in a previous study (RTOG 84–01). Sixty six patients at six institutions were enrolled on a prospective Phase I/II study. Eligible deep seated tumours were treated with a combination of external hyperthermia and radiation therapy. Radiation consisted of 1·7–2 Gy per fraction, 4–5 fractions per week, to > 20 Gy (previously irradiated lesions) or > 50 Gy (no previous radiation). Deep hyperthermia was delivered with electromagnetic devices: BSD 2000 for 92% of cases, Thermotron for 5% of cases, other low frequency electromagnetic for 4% of cases. Hyperthermia was delivered ≤ twice weekly. Overall complete and partial response rates were 34% and 16% respectively. Response was not correlated with maximum tumour temperature or disease site. There was, however, a strong association with radiation dose: 54% CR with ≥ 45 Gy versus 7% with < 45 Gy (p < 0·0001). The achieved temperatures were less than ideal. Although the average maximum tumor temperature was 41·9°C (range 35·7°C–46·7°C), the minimum tumour temperatures were low. The average minimum tumour temperature was 38·5°C and was never > 41·8°C. Treatment was well tolerated with no fatalities. There were four acute grade 3 or 4 toxicities (6% of patients). Patient discomfort resulted in interruption or discontinuation of sessions in 30% of the sessions. In 12 cases (18% of patients) the planned course of hyperthermia was discontinued due to acute discomfort. The devices used in this study were better tolerated than the devices used in the previous Phase I/II deep hyperthermia trial (RTOG 84·01) with less patient discomfort and no problems with severe systemic cardiovascular stress. In the previous study 68% of the hyperthermia courses were prematurely terminated primarily due to patient discomfort and toxicity; in the present study 18% were prematurely terminated. However, as indicated by the low minimum tumour temperature, fundamental problems with achieving acceptable temperature distributions remain.     

Alternating radiotherapy and chemotherapy for inoperable stage III non–small-cell lung cancer: long-term results of two phase II GOTHA trials

Purpose/Objective: To report on two consecutive Phase II cooperative trials in which we evaluated the combination of alternating hyperfractionated accelerated radiotherapy and cisplatin-based chemotherapy in inoperable Stage III non-small cell lung cancer (NSCLC).Patients & Methods: Between February 1986 and September 1989, 65 patients were entered in the first trial (GOTHA I), and between December 1989 and October 1992 67 were enrolled in the second trial (GOTHA II). In both protocols, radiotherapy (RT) was administered twice daily, at 6 h intervals, 5 days a week, to a total dose of 63 Gy in 42 fractions of 1.5 Gy. RT was given during weeks 2, 3, 6, and 7, over an elapsed time of 6 weeks. In GOTHA I, three cycles of cisplatin, 60 mg/m² day 1, mitomycin, 8 mg/m² day 1, and vindesin 3 mg/m² day 1 and the first day of the following week, were given during weeks 1, 5, and 9; in GOTHA II, cisplatin 70 mg/m² day 1 and vinblastin 5 mg/m² day 1 and the first day of the following week were given during weeks 1, 5, 9, 13, 17, and 21.Results: With a minimum follow-up of 3 years, the 1-, 2-, 5-, and 8-year overall survival probability was 56% (95% CI 47–64%), 27% (20–35%), 12% (7–18%) and 9% (3–16%), respectively, with a median survival of 13.6 months (11.4–16.8). Median follow-up for survivors was 6 years (3.3–9.9). There were no survival differences between Stages IIIA and IIIB (p = 0.84), performance status 0, 1, 2 (p = 0.87), sex (p = 0.45) or between the two treatment protocols. At this time, 14 patients are alive, and 118 have died: 102 from NSCLC, 4 from acute toxicity, 2 from secondary surgery, 4 from other medical causes, and 6 from unknown causes. Correlation between response and long-term survival was poor, since of the 24 patients who survived 3 years or more, only 6 (25%) were classified as having a complete response; the remainder having either a partial response (11, 46%), no change (6, 25%), or “progressive disease” (1, 4%). First site of relapse was local in 31% of these cases, distant in 43%, local and distant in 15%, and unknown in 11%. Main grade 3–4 acute toxicities were nausea-vomiting (17%), mucositis (15%), leukopenia (41%), and thrombocytopenia (11%). Eight patients presented with grade 3–4 symptomatic lung radiation pneumopathy.Conclusion: Based on this experience with 132 patients, this combination of alternated RT and chemotherapy (CT) for inoperable Stage III NSCLC is feasible with acceptable toxicity, and long-term results suggest a gain in survival when compared to those obtained with conventional RT alone. However, the still high local and distant failure rates indicate that both local and systemic therapies need to be improved.     

Patterns of nodal relapse after surgery and postoperative radiation therapy for carcinomas of the major and minor salivary glands: What is the role of elective neck irradiation?

PURPOSE: To evaluate the incidence of nodal relapses from carcinomas of the salivary glands among patients with clinically negative necks in an attempt to determine the potential utility of elective neck irradiation (ENI). METHODS AND MATERIALS: Between 1960 and 2004, 251 patients with clinically N0 carcinomas of the salivary glands were treated with surgery and postoperative radiation therapy. None of the patients had undergone previous neck dissection. Histology was: adenoid cystic (84 patients), mucoepidermoid (60 patients), adenocarcinoma (58 patients), acinic cell (21 patients), undifferentiated (11 patients), carcinoma ex pleomorphic adenoma (7 patients), squamous cell (7 patients), and salivary duct carcinoma (3 patients); 131 patients (52%) had ENI. Median follow-up was 62 months (range, 3-267 months). RESULTS: The 5- and 10-year actuarial estimates of nodal relapse were 11% and 13%, respectively. The 10-year actuarial rates of nodal failure were 7%, 5%, 12%, and 16%, for patients with T1, T2, T3, and T4 disease, respectively (p = 0.11). The use of ENI reduced the 10-year nodal failure rate from 26% to 0% (p = 0.0001). The highest crude rates of nodal relapse among those treated without ENI were found in patients with squamous cell carcinoma (67%), undifferentiated carcinoma (50%), adenocarcinoma (34%), and mucoepidermoid carcinoma (29%). There were no nodal failures observed among patients with adenoid cystic or acinic cell histology. CONCLUSION: ENI effectively prevents nodal relapses and should be used for select patients at high risk for regional failure.     

Phase I–II study of plitidepsin and dacarbazine as first-line therapy for advanced melanoma

Background:

This phase I–II trial compared plitidepsin 1-h infusion alone or combined with dacarbazine (DTIC) 1-h infusion as front-line therapy for advanced melanoma.

Methods:

The recommended dose (RD) for plitidepsin/DTIC was defined in the first stage. In the second stage, patients were randomised to receive single-agent plitidepsin 3.2 mg m⁻² (n=20) on days 1, 8 and 15 every 4 weeks (q4wk) or plitidepsin 2.4 mg m⁻² on days 1, 8 and 15 q4wk combined with DTIC 800 mg m⁻² q4wk (n=38).

Results:

The overall response rate with plitidepsin/DTIC was 21.4% all responders had normal serum lactate dehydrogenase (LDH) levels and performance status ⩽1 at baseline. Median progression-free survival (PFS) with plitidepsin/DTIC was 3.3 months in all patients, and 4.3 months in those with baseline normal LDH. No responses occurred with single-agent plitidepsin and median PFS was 1.5 months. Both regimens were well tolerated. Haematological abnormalities were more common and transaminase increases more severe with plitidepsin/DTIC. Treatment-related transaminase increases leading to infusion omission on day 8 were relatively common. No drug–drug pharmacokinetic interactions were found.

Conclusion:

This plitidepsin/DTIC schedule has antitumour activity and manageable toxicity in advanced melanoma. Further evaluation of plitidepsin 2.4 mg m⁻² fortnightly and DTIC 800 mg m⁻² q4wk is recommended.     

Histologic factors in the grading and prognosis of astrocytoma grade I–IV

Summary The prognostic value of histologic or cytologic features were examined in 317 patients with an astrocytic glioma. Of 46 features examined 7 morphologic characteristics of nucleus and 5 of mesenchyma correlated well with grading according to Kernohan. The morphological characteristics of nucleus are cellularity, atypical nuclei, polymorphism, multinucleated cells, hyperchromasia, gigantic nuclei and mitosis. For mesenchyma these were vascularity, vascular endothelial proliferation, vascular glomeruli, necrosis with palisade formation and necrosis without palisade formation. Scoring of these items on a scale from 0 to 4 enables us to establish a nucleus-score (max. 28) and a mesenchyma-score (max. 20). Nucleus-score + mesenchyma-score 10 corresponds with astrocytoma grade II, 20 with grade III and > 20 with grade IV. A low nucleus-score in grade II (< 5) predicts a long survival (> 10 years), a high nucleus-score a low survival (median 0.6 year). In grade III a low mesenchyma-score predicts a median survival of 1.25 years, a high mesenchyma-score a median survival of 0.38 year.This retrospective study shows that nucleus-score and mesenchyma-score correlate to grading according to Kernohan and are highly correlative to survival.