Advances in radiotherapy delivery for rectal cancer: a European perspective.

Preoperative chemoradiotherapy and radiotherapy with an integrated boost offer excellent local control rates in patients with rectal cancer. The introduction of intensity-modulated radiotherapy and image-guided radiotherapy has drastically improved the tolerance of these treatments. The new challenge is developing organ-preserving strategies and curative treatments for medically inoperable patients. Contact radiotherapy seems efficient for small tumors. Tumor hypoxia limits the success of radiotherapy for locally advanced cancers. Modulation of the L-arginine/iNOS pathway and implementation of hypoxia imaging in radiotherapy planning may overcome this hurdle.

Advances in radiotherapy and targeted therapies for rectal cancer.

The last decade witnessed a significant progress in understanding the biology and immunology of colorectal cancer alongside with the technical innovations in radiotherapy. The stepwise implementation of intensity-modulated and image-guided radiation therapy by means of megavolt computed tomography and helical tomotherapy enabled us to anatomically sculpt dose delivery, reducing treatment related toxicity. In addition, the administration of a simultaneous integrated boost offers excellent local control rates. The novel challenge is the development of treatment strategies for medically inoperable patient and organ preserving approaches. However, distant control remains unsatisfactory and indicates an urgent need for biomarkers that predict the risk of tumor spread. The expected benefit of targeted therapies that exploit the tumor genome alone is so far hindered by high cost techniques and pharmaceuticals, hence hardly justifying rather modest improvements in patient outcomes. On the other hand, the immune landscape of colorectal cancer is now better clarified with regard to the immunosuppressive network that promotes immune escape. Both N2 neutrophils and myeloid-derived suppressor cells (MDSC) emerge as useful clinical biomarkers of poor prognosis, while the growing list of anti-MDSC agents shows promising ability to boost antitumor T-cell immunity in preclinical settings. Therefore, integration of genetic and immune biomarkers is the next logical step towards effective targeted therapies in the context of personalized cancer treatment.

Preoperative intensity-modulated and image-guided radiotherapy with a simultaneous integrated boost in locally advanced rectal cancer: report on late toxicity and outcome.

BACKGROUND AND PURPOSE: The addition of chemotherapy to preoperative radiotherapy has been established as the standard of care for patients with cT3-4 rectal cancer. As an alternative strategy, we explored intensity-modulated and image-guided radiotherapy (IMRT-IGRT) with a simultaneous integrated boost (SIB) in a prospective phase II study. Here, we report outcome and late toxicity after a median follow-up of 54 months. METHODS AND MATERIALS: A total of 108 patients were treated preoperatively with IMRT-IGRT, delivering a dose of 46 Gy in fractions of 2 Gy. Patients (n=57) displaying an anticipated circumferential resection margin (CRM) of less than 2mm based on magnetic resonance imaging received a SIB to the tumor up to a total dose of 55.2 Gy. RESULTS: The absolute incidence of grade ≥3 late gastrointestinal and urinary toxicity was 9% and 4%, respectively, with a 13% rate of any grade ≥3 late toxicity. The actuarial 5-year local control (LC), progression-free survival (PFS) and overall survival (OS) were 97%, 57%, and 68%. On multivariate analysis, R1 resection and pN2 disease were associated with significantly impaired OS. CONCLUSIONS: The use of preoperative IMRT-IGRT with a SIB resulted in a high 5-year LC rate and non-negligible late toxicity.

Hypoxia integration in the serological proteome analysis unmasks tumor antigens and fosters the identification of anti-phospho-eEF2 antibodies as potential cancer biomarkers.

The expression by tumor cells of proteins with aberrant structure, expression or distribution accounts for the development of a humoral immune response. Autoantibodies (aAb) directed against tumor-associated antigens (TAA) may thus be particularly relevant for early detection of cancer. Serological proteome analysis (SERPA) aims to identify such circulating aAb through the immunoblotting of 2D-separated tumor cell proteins with cancer patient serum and the consecutive MS identification of proteins in reactive spots. This method has the advantage to use post-translationally modified proteins as a source of potential TAA. Here, we applied this strategy by using colorectal tumor cells pre-exposed to hypoxia in order to promote the expression of a pattern of TAA more likely to represent in vivo conditions. We used two human HCT116 and HT29 colorectal cancer cell lines exposed for 48 hours to 1% O2. Spots positive after immunoblotting of 2D-separated lysates of hypoxic cells with the sera of tumor-bearing mice, were collected and analysed by MS for protein identification. Among the hypoxia-specific immunogenic proteins, we identified a phosphorylated form of eukaryotic translation elongation factor 2 (phospho-Thr56 eEF2). We confirmed the increased phosphorylation of this protein in hypoxic colorectal tumor cells as well as in mouse tumors. Using a specific immunoassay, we could detect the presence of corresponding anti-phospho-Thr56 eEF2 aAb in the serum of tumor-bearing mice (vs healthy mice). We further documented that the detection of these aAb preceded the detection of a palpable tumor mass in mice and validated the presence of anti-phospho-Thr56 eEF2 aAb in the serum of patients with adenomatous polyps and colorectal carcinoma. In conclusion, this study validates a phosphorylated form of eEF2 as a new TAA and more generally, provides evidence that integrating hypoxia upstream of SERPA offers a more relevant repertoire of TAA able to unmask the presence of circulating aAb.

Hepatocytes determine the hypoxic microenvironment and radiosensitivity of colorectal cancer cells through production of nitric oxide that targets mitochondrial respiration.

PURPOSE: To determine whether host hepatocytes may reverse hypoxic radioresistance through nitric oxide (NO)-induced oxygen sparing, in a model relevant to colorectal cancer (CRC) liver metastases. METHODS AND MATERIALS: Hepatocytes and a panel of CRC cells were incubated in a tissue-mimetic coculture system with diffusion-limited oxygenation, and oxygen levels were monitored by an oxygen-sensing fluorescence probe. To activate endogenous NO production, cocultures were exposed to a cytokine mixture, and the expression of inducible nitric oxide synthase was analyzed by reverse transcription-polymerase chain reaction, Western blotting, and NO/nitrite production. The mitochondrial targets of NO were examined by enzymatic activity. To assess hypoxic radioresponse, cocultures were irradiated and reseeded for colonies. RESULTS: Resting hepatocytes consumed 10-40 times more oxygen than mouse CT26 and human DLD-1, HT29, HCT116, and SW480 CRC cells, and thus seemed to be the major effectors of hypoxic conditioning. As a result, hepatocytes caused uniform radioprotection of tumor cells at a 1:1 ratio. Conversely, NO-producing hepatocytes radiosensitized all CRC cell lines more than 1.5-fold, similar to the effect of selective mitochondrial inhibitors. The radiosensitizing effect was associated with a respiratory self-arrest of hepatocytes at the level of aconitase and complex II, which resulted in profound reoxygenation of tumor cells through oxygen sparing. Nitric oxide-producing hepatocytes were at least 10 times more active than NO-producing macrophages to reverse hypoxia-induced radioresistance. CONCLUSIONS: Hepatocytes were the major determinants of the hypoxic microenvironment and radioresponse of CRC cells in our model of metabolic hypoxia. We provide evidence that reoxygenation and radiosensitization of hypoxic CRC cells can be achieved through oxygen sparing induced by endogenous NO production in host hepatocytes.

Current status of intensified neo-adjuvant systemic therapy in locally advanced rectal cancer.

The addition of 5-fluorouracil (5-FU) or its prodrug capecitabine to radiotherapy (RT) is a standard approach in the neo-adjuvant treatment of patients with rectal tumors extending beyond the muscularis propria (stage II) and/or with clinical evidence of regional lymph node metastases (stage III). According to European randomized trials, the combined treatment modality resulted in favorable local control rates as compared with radiotherapy (RT) alone, but no improvement was found regarding the occurrence of distant metastases or overall survival. In an effort to further enhance the response rates and to decrease the high incidence of distant metastases in locally advanced rectal cancer patients, the addition of other chemotherapeutical drugs and biologic agents as radiation sensitizers to neo-adjuvant 5-FU based chemoradiotherapy (CRT) has been recently investigated. The role of those agents is however questionable as first results from phase III data do not show improvement on pathologic complete remission and circumferential resection margin negative resection rates as compared to 5-FU based CRT, nevertheless an increased toxicity.

Phase II study of helical tomotherapy in the multidisciplinary treatment of oligometastatic colorectal cancer.

BACKGROUND: Complete metastasectomy provides a real chance for long-term survival in patients with oligometastatic colorectal cancer (CRC). For inoperable patients, we evaluated in this study intensity-modulated and image-guided radiotherapy (IMRT-IGRT) by helical tomotherapy. METHODS: Twenty-four CRC patients with ≤ 5 metastases were enrolled, receiving a dose of 50 Gy in fractions of 5 Gy. No limitations concerning dimension or localization of the metastases were imposed. Whole body PET-CT was performed at baseline and 3 months after the initiation of RT to evaluate the metabolic response rate according to PET Response Criteria in Solid Tumors (PERCIST) version 1.0. RESULTS: A total of 53 metastases were treated. Seventeen patients (71%) received previously ≥ 1 line of chemotherapy for metastatic disease, displaying residual (n = 7) or progressive (n = 10) metabolic active oligometastatic disease at time of inclusion. Most common sites were the lung, liver and lymphnodes. One patient (4%) experienced grade 3 dysphagia. Twenty-two patients were evaluated by post-treatment PET-CT. Twelve patients achieved a complete (n = 6) or partial (n = 6) metabolic response, resulting in an overall metabolic response rate of 55%. At a median follow-up of 10 months, 7 patients (29%) are in remission, of which 5 received previous chemotherapy with residual oligometastatic disease at time of inclusion. The actuarial 1-year local control, progression-free survival, and overall survival were 54%, 14% and 78%. CONCLUSIONS: Helical tomotherapy delivering 10 fractions of 5 Gy resulted in a metabolic response rate of 55%, and appeared to be attractive as consolidation of inoperable oligometastatic disease after effective chemotherapy. TRIAL REGISTRATION: Eudract 2008-008300-40; NCT00807313.

Phase II study of preoperative helical tomotherapy with a simultaneous integrated boost for rectal cancer.

PURPOSE: The addition of concomitant chemotherapy to preoperative radiotherapy is considered the standard of care for patients with cT3-4 rectal cancer. The combined treatment modality increases the complete response rate and local control (LC), but has no impact on survival or the incidence of distant metastases. In addition, it is associated with considerable toxicity. As an alternative strategy, we explored prospectively, preoperative helical tomotherapy with a simultaneous integrated boost (SIB). METHODS AND MATERIALS: A total of 108 patients were treated with intensity-modulated and image-guided radiotherapy using the Tomotherapy Hi-Art II system. A dose of 46 Gy, in daily fractions of 2 Gy, was delivered to the mesorectum and draining lymph nodes, without concomitant chemotherapy. Patients with an anticipated circumferential resection margin (CRM) of less than 2 mm, based on magnetic resonance imaging, received a SIB to the tumor up to a total dose of 55.2 Gy. Acute and late side effects were scored using the National Cancer Institute Common Terminology Criteria for Adverse Events version 3.0. RESULTS: A total of 102 patients presented with cT3-4 tumors; 57 patients entered the boost group and 51 the no-boost group. One patient in the no-boost group developed a radio-hypersensitivity reaction, resulting in a complete tumor remission, a Grade 3 acute and Grade 5 late enteritis. No other Grade ≥3 acute toxicities occurred. With a median follow-up of 32 months, Grade ≥3 late gastrointestinal and urinary toxicity were observed in 6% and 4% of the patients, respectively. The actuarial 2-year LC, progression-free survival and overall survival were 98%, 79%, and 93%. CONCLUSIONS: Preoperative helical tomotherapy displays a favorable acute toxicity profile in patients with cT3-4 rectal cancer. A SIB can be safely administered in patients with a narrow CRM and resulted in a promising LC.

Prediction of response to neoadjuvant radiotherapy in patients with locally advanced rectal cancer by means of sequential 18FDG-PET.

PURPOSE: Morphologic imaging techniques perform poorly in assessing the response to preoperative radiotherapy (RT), mainly because of desmoplastic reactions. The aim of this study was to investigate the potential of sequential 18-fluoro-2-deoxy-d-glucose (18FDG-PET) in assessing the response of rectal cancer to neoadjuvant RT and to determine which parameters can be used as surrogate markers for histopathologic response. METHODS AND MATERIALS: 18FDG-PET scans were acquired before and during the 5th week after the end of RT. Tracer uptake was assessed semiquantitatively using standardized uptake values (SUV). The percentage differences (%Δ) between pre- and post-RT scans in SUV(max), SUV(mean), metabolic volume (MV), and total glycolytic volume (tGV) were calculated. RESULTS: Forty-five consecutive patients with histologically confirmed rectal adenocarcinoma were enrolled. After neoadjuvant RT, 20 of the 45 patients were classified as histopathologic responders and 25 as non-responders. Intense 18F-FDG uptake was seen in all tumors before neoadjuvant RT (average SUV(max) 12.9 ± 6.0). When patients were classified as histologic responders and nonresponders, significant differences in %ΔSUV(max) (55.8% vs. 37.4%, p = 0.023) and %ΔSUV(mean) (40.1% vs. 21.0%, p = 0.001) were observed between the two groups. For %ΔMV and %ΔtGV, decreases were more prominent in responders but were not significantly different from those in nonresponders. As demonstrated by receiver operating characteristic analysis, %ΔSUV(mean) was a more powerful discriminator than was %ΔSUV(max). The sensitivity, specificity, accuracy, positive predictive value, and negative predictive value for optimal threshold of %ΔSUV(mean) (24.5%) were 80%, 72%, 76%, 70%, and 82% respectively. CONCLUSION: Sequential 18FDG-PET allows assessment of the response to preoperative RT. Both %ΔSUV(mean) and %ΔSUV(max) correlate with histopathologic response and can be used to evaluate and compare the effectiveness of different neoadjuvant treatment strategies. The maximum accuracy figures and the positive predictive value figures for both Δ%SUV(mean) and Δ%SUV(max) are, however, too low to justify modification of the standard treatment protocol of an individual patient.

Preoperative helical tomotherapy and megavoltage computed tomography for rectal cancer: impact on the irradiated volume of small bowel.

PURPOSE: Preoperative (chemo)radiotherapy is considered to be standard of care in locally advanced rectal cancer, but is associated with significant small-bowel toxicity. The aim of this study was to explore to what extent helical tomotherapy and daily megavolt (MV) CT imaging may reduce the irradiated volume of small bowel. METHODS AND MATERIALS: A 3D-conformal radiotherapy (3D-CRT) plan with CTV-PTV margins adjusted for laser-skin marks (15, 15, and 10 mm for X, Y, and Z directions, respectively) was compared with helical tomotherapy (IMRT) using the same CTV-PTV margins, and to helical tomotherapy with margins adapted to daily MV-CT imaging (IMRT/IGRT; 8, 11, 7, and 10 mm for X, Y(ant), Y(post) and Z resp.) for 11 consecutive patients. The planning goals were to prescribe 43.7 Gy to 95% of the PTV, while minimizing the volume of small bowel receiving more than 15 Gy (V(15 SB)). RESULTS: The mean PTV was reduced from 1857.4 +/- 256.6 cc to 1462.0 +/- 222.3 cc, when the CTV-PTV margins were adapted from laser-skin marks to daily MV-CT imaging (p < 0.01). The V(15 SB) decreased from 160.7 +/- 102.9 cc to 110.9 +/- 74.0 cc with IMRT and to 81.4 +/- 53.9 cc with IMRT/IGRT (p < 0.01). The normal tissue complication probability (NTCP) for developing Grade 2+ diarrhea was reduced from 39.5% to 26.5% with IMRT and to 18.0% with IMRT/IGRT (p < 0.01). CONCLUSION: The combination of helical tomotherapy and daily MV-CT imaging significantly decreases the irradiated volume of small bowel and its NTCP.

Lipid a radiosensitizes hypoxic EMT-6 tumor cells: role of the NF-kappaB signaling pathway.

PURPOSE: Lipid A has shown promising immunostimulatory effects in both experimental tumor models and advanced stage cancer patients. This study examines whether lipid A may directly modulate the radioresponse of tumor cells by activating inducible nitric oxide synthase (iNOS) or cyclooxygenase-2 (COX-2) through nuclear factor-kappaB (NF-kappaB) signaling. METHODS AND MATERIALS: Hypoxic EMT-6 tumor cells were exposed to lipid A and analyzed for the level of COX-2 and iNOS by Western blotting and enzymatic assays. The hypoxic radioresponse of EMT-6 cells was estimated by clonogenic survival. The activation of NF-kappaB was examined by immunostaining of its p65 subunit and by luciferase reporter gene assay. RESULTS: Lipid A dose-dependently increased the expression and activity of iNOS with a maximal effect at plasma achievable concentrations of 3-30 micro g/mL. The COX-2 mediated production of prostaglandin E2 was constitutively high and further upregulated by lipid A. The radiosensitivity of hypoxic EMT-6 cells was increased up to 2.5 times and counteracted by the iNOS inhibitor aminoguanidine but not by the COX-2 inhibitor NS-398. The mechanism of radiosensitization was linked to NF-kappaB signaling, because its inhibition by phenylarsine oxide impaired both iNOS activation and radioresponse. CONCLUSION: Lipid A is an efficient hypoxic cell radiosensitizer at plasma relevant concentrations, which provides a rationale to combine lipid A with radiotherapy in further studies.

Metastases to the thyroid gland-a report of six cases.

The association of an asymptomatic mass, normal thyroid function, and a cold nodule can occur months to years after a primary cancer. Work-up should include ruling out other metastases and fine-needle aspiration cytology. We report six cases of secondary thyroid cancer. Two of the patients in our series presented with hyperthyroidism, which may be due to invasion and disruption of thyroid follicles.