2. Results
2.1. Patients and Treatment Characteristics
A cohort of 987 consecutive non-metastatic AC patients treated between 2007–2019 was analyzed. Patients were enrolled within 25 different Italian centers. Clinical characteristics for all treated patients are shown in
Table 1.
An Eastern Cooperative Oncology Group (ECOG) performance status (PS) of 0 was reported in 775 patients (78.5%). HIV positivity was identified in 90 (9.1%) of the evaluated patients. In most of the patients, HPV status was not reported (55.5%). Tumors were located within the anal canal in 879 patients (89.1%). Basaloid histology was described in 84 patients (8.5%).
Three hundred thirty patients (33.4%) had cT1-2N0 stage, 106 patients (10.6%) had cT3-4N0 stage and 551 (55.8%) had lymph node involvement. Overall, 633 patients (64.1%) presented with locally advanced disease (LAD) at diagnosis. Twenty-four patients (2.4%) had a single site of clinical abdominal lymph node involvement (lumbar–aortic and/or common iliac lymph nodes).
Imaging modalities used to define tumor (T), nodal (N) and distant metastases (M) stages are shown in
Table 2.
Magnetic resonance imaging (MRI) was used in 541 patients (54.8%) to define T stage; 18FDG-PET was used in 375 patients (38.0%) and 467 patients (47.3%) for N and M stage identification, respectively.
Treatment characteristics are detailed in
Table 3.
Three hundred and two patients (30.6%) were treated with static or dynamic IMRT, 470 patients (47.6%) with volumetric modulated arc therapy (VMAT) and 215 patients (21.8%) with tomotherapy. A SIB strategy was used in 568 patients (57.5%). An additional sequential boost was administered in 167 patients (16.9%) using external beam RT (EBRT) or brachytherapy (BRT) in 122 patients (12.4%) and 45 patients (4.6%), respectively. Concomitant chemotherapy was administered in 934 patients (94.63%). The mitomycin (MMC) plus 5-fluorouracil (5-FU) or capecitabine regimen was administered in 779 patients (78.92%).
The total RT dose and daily fractionation prescription were analyzed according to clinical stage at presentation and reported based on the low-, intermediate- and high-risk planning target volumes (PTVs), as defined in the RTOG 0529 study [
15]. The median total dose was 55 Gy (range: 45–75).
In patients with cT1-2 N0 disease, the elective low-risk PTV received a median dose of 45 Gy (range 32.40–48 Gy, 1.40–2 Gy daily) and high-risk PTV received a median dose of 54 Gy (range 37.50–70.40 Gy, 1.80–2.40 Gy daily), with a median total dose of 55 Gy (range 46–70.40 Gy).
In patients with cT3-T4 N0 disease, the elective low-risk PTV received a median dose of 45 Gy (range 34.50–48 Gy, 1.40–1.80 Gy daily) and high-risk PTV received a median dose of 54 Gy (range 41.40–68.40 Gy, 1.80–2.40 Gy daily), with a median total dose of 55 Gy (range 50–73 Gy).
In patients with cT1-T2 N+ disease, the elective low-risk PTV received a median dose of 45 Gy (range 35.8–54 Gy, 1.40–2 Gy daily) and high-risk PTV received a median dose of 54 Gy (range 40–68.4 Gy, 1.80–2.40 Gy daily), with a median total dose of 55 Gy (range 45–74.4 Gy).
In patients with cT3-T4 N+ disease, the elective low-risk PTV received a median dose of 45 Gy (range 36–54 Gy, 1.40–2 Gy daily) and high-risk PTV received a median dose of 55 Gy (range 44–72.4 Gy, 1.80–2.40 Gy daily), with a median total dose of 56 Gy (range 45–74.8 Gy).
2.2. Treatment Compliance and Toxicity
The median OTT was 45 days (range: 25–115). A treatment interruption of >5 days for toxicity occurred in 186 patients (18.8%). RT was not definitively completed in 67 patients (6.8%). Detailed toxicity profiles are shown in
Table 4.
The most common acute toxicities were grade 2 dermatitis, reported in 491 patients (49.75%), and grade 1 diarrhea, reported in 371 patients (37.6%). Severe (grade > 3) acute toxicity was observed mainly as skin toxicity (desquamation) in 253 patients (25.6%). Other severe toxicities were GI in 65 patients (6.6%) and urogenital in 5 patients (0.5%). Overall, the grade 3–4 acute toxicity rate was 32.7%. The acute hematologic toxicity was reported in 265 patients (26.8%) as grade 1, in 126 patients (12.8%) as grade 2 and in 86 patients (8.7%) as grade >3 toxicity.
With a median follow-up of 28 months (range 6–138), GI late effects (diarrhea and proctitis) were the most common late toxicities: grade 1 in 205 patients (20.8%), grade 2 in 65 patients (6.6%) and grade 3–4 in 18 patients (1.8%). Grade 1, 2 and 3–4 late skin toxicities were observed in 183 patients (18.5%), 10 patients (1%) and 2 patients (0.2%), respectively. Pelvic bone fractures or density alterations were reported in 16 patients (1.6%). Sexual disorders and dyspareunia occurred in 9 (0.9%) and 3 (0.3%) female patients, respectively. Late anemia and/or thrombocytopenia was described in 5 patients (0.5%).
2.3. Treatment Response and Clinical Outcomes
The diagnostic imaging modalities allowing for tumor response evaluation were MRI in 386 patients (39.10%), CT in 184 patients (18.65%), anoscopy in 146 patients (14.80%), endorectal ultrasound (ERUS) in 98 patients (10%) and 18FDG-PET in 71 patients (7.2%). Multiple diagnostic imaging modalities were used in combinations in 90 patients (9.12%).
At 3 months after the start of RT, 536 patients (54.3%) had complete response (CR), 291 patients (29.5%) partial response (PR), 12 patients (1.2%) stable disease (SD), and 13 patients (1.3%) had local progressive disease (PD). For 135 patients (13.7%), data were not available at the 3-month analysis.
At 6 months after the start of RT, 766 patients (77.7%) had CR, 140 patients (14.2%) had PR, 17 patients (1.7%) had SD, and 57 patients (5.8%) had PD. Data were not available at the 6-month analysis for 7 patients (0.7%). The overall clinical CR rate (beyond 6-month evaluations) was 90.6%.
The 2- and 3-year LC rates were 86.9% (95% CI: 85.6–88.2%) and 85.8% (95% CI: 84.4–87.2%), respectively (
Figure 1a).
A total of 84 patients underwent a colostomy with abdominoperineal resection in the first year after the start of RT, with an estimated cumulative incidence of colostomy at 12 months of 9.3% (95% CI: 8.2–10.3%). The 2- and 3-year colostomy-free survival (CFS) rates were 81.5% (95% CI: 79.83–83.1%) and 77.9% (95% CI: 76.1–79.8%), respectively (
Figure 1a).
The 2-, 3- and 5-year overall survival (OS) rates were 92.3% (95% CI: 91.3–93.3%), 88.1% (95% CI: 78.8–89.4%) and 82.9% (95% CI: 81.0–84.7%), with 2- and 3-year progression-free survival (PFS) rates of 83.2% (95% CI: 81.6–84.7%) and 80.2% (95% CI: 78.5–82.0%), respectively (
Figure 1b). Overall, the 2-, 3- and 5-year event-free survival (EFS) rates were 76.6% (95% CI: 73.8–79.4%), 73.4% (95% CI: 70.5–76.5%) and 69.4% (95% CI: 66.0–73.0%), respectively.
2.4. Univariate and Multivariate Analyses
As study variables, we evaluated age, gender, ECOG PS, HPV and HIV status, clinical stage (T1-T2 vs. T3-T4), lymph node involvement (N0 vs. N+), histological grade (G1-G2 vs. G3), and treatment parameters, such as total dose and treatment duration (OTT: <45 vs. ≥45 days).
Univariate analysis assessing overall clinical CR (after 6 months) showed that histological grade 3 was associated with a low probability of CR (odds ratio (OR) 0.11, 95% CI 0.03–0.52, p = 0.030). In addition, node-positive patients had a significantly lower probability of CR than patients with uninvolved lymph nodes (OR 0.45, 95% CI 0.25–0.76, p = 0.004). Finally, a significant association with disease extension was reported (OR 0.10, 95% CI 0.04–0.27, p < 0.001 and OR = 0.50, 95% CI 0.28–0.85, p = 0.013 for extended disease (ED) and LAD, respectively).
As shown in
Table 5 on univariate analysis, a statistically significant lower CFS, OS, PFS and EFS was found for patients with poor ECOG PS and an advanced stage. Lymph node involvement (regardless of T-stage) and histological grade 3 were associated with a lower LC rate (
p < 0.001 and
p = 0.025, respectively). Age >68.5 (cut-off set at 68.5 years as median age of population study) showed a trend for a lower PFS (
p = 0.052). Moreover, the correlation between clinical outcomes and disease extension is shown in
Figure 2a,b.
No statistically significant association was found between RT total dose and/or boost modality (SIB) and clinical outcomes. OTT > 45 days showed a trend with a lower PFS (
p = 0.050) and was significantly associated with a lower EFS (
p = 0.030) (
Table 6).
According to the multivariate analysis, lymph node involvement negatively affected all clinical outcome measures (LC, CFS, OS, PFS and EFS). Age > 68.5 (cut-off set at 68.5 years as median age of population study) and pathological grade 3 were confirmed as negative prognostic factors for PFS (p = 0.019) and LC (p = 0.032), respectively.
Investigating the relationships between toxicity (all grades) and the clinical and dosimetric parameters due to the high dimensionality of the comparisons, we calculated ORs and CIs (95%) considering a significant p-value of p < 0.001. In particular, disease extension results were associated with acute skin toxicity (ED vs. Early: OR = 4.84, 95% CI 2.13–12.04; LAD vs. Early: OR = 0.60, 95% CI 0.46–0. 76), acute gastrointestinal toxicity (LAD vs. Early: OR = 0.51, 95% CI 0.4–0.65) and with acute urological toxicity (LAD vs. Early: OR = 0.36, 95% CI 0.27–0.46). Disease extension results were also associated with skin (ED vs Early: OR = 19.64, 95% CI 8.3–48.63) and subcutaneous late toxicity (LAD vs. Early: OR = 0.43, 95% CI 0.3–0.61).
3. Discussion
CRT provided high rates of complete responders (up to 80–90%) in several trials [
1,
2,
3,
4,
5,
6,
7,
8,
9] and currently represents the standard treatment for stage II–III AC patients. Radiotherapy delivery in anal cancer is complex because of the target shape and the proximity to dose-sensitive organs at risk (OARs), such as bowel, femoral heads, bladder, genitalia and perineal skin. IMRT techniques, modulating the beam fluence during delivery, offer the possibility of a dose-painted treatment with high doses to the tumor, minimizing the dose to surrounding OARs. This potentially allows the reduction of acute and late adverse events, improving the therapeutic ratio. The consequent decrease in acute toxicity improves treatment compliance, with a decrease in interruptions and a shorter OTT, with a potential beneficial effect on clinical outcomes. Several studies have shown promising results using this approach [
12,
13,
14,
15,
16,
17,
18,
19,
20,
21,
22], and it should be considered the standard of care for anal cancer [
23].
In our study, all patients were treated with IMRT techniques.
We observed a substantial clinical response rate: 6-moCR of 77.7% and an overall clinical CR rate (beyond 6-month evaluations) of 90.6%, in agreement with previous studies.
The ACT II trial reported a 26-week CR rate of 89.6% in the cisplatin group and 90.5% in the MMC group, showing that patients with cCR at 26 weeks had a superior 5-year OS compared to patients without cCR [
7]. Based on these data, the European Society for Medical Oncology (ESMO) guidelines recommend a clinical evaluation for CR and endorsed a watchful wait approach, stating that partial regression can be managed by a close follow-up to confirm that complete regression occurs within 6 months [
23].
Five-year OS in patients with stage II-III anal squamous cell cancer, treated with CRT, is approximately 75%. RTOG 9811 and ACT II used conformal RT techniques with concurrent 5-FU and MMC and reported 5-year OS rates of 78.3% and 79.0%, respectively [
6,
8,
9]. Long term results (median follow-up of 49 months) of a single institution study of 99 patients, treated with dose-painted IMRT according to the RTOG 0529 trial, showed a 4-year OS of 85.8% [
17]. Moreover, in an Italian mono-institutional cohort of 87 patients treated with SIB-IMRT, the 3-year OS rate was 79%, with a 3-year CFS and LC rate of 64% and 69%, respectively [
18]. In our study, the estimated 3- and 5-year OS rates were 88.09% (95% CI: 78.76–89.42) and 82.86% (95% CI: 81–84.72), respectively. Consistent results in terms of 3-year LC (85.84%; 95% CI: 84.43–87.25), 3-year CFS (77.94%; 95% CI: 76.07–79.81) and 3-year PFS (80.24; 95% CI: 78.5–81.98) rates were also observed.
Several trials reported that high conformal techniques, such as IMRT, VMAT and tomotherapy, require an accurate definition and delineation of treatment volumes to maintain or potentially improve locoregional control. In AC patients, the primary tumor site, pelvic and inguinal nodes are the main areas of recurrence and could represent three different targets for RT planning. Recommendations based on expert views concerning the primary tumor and elective lymph nodes delineation have been provided [
24,
25]. On the other hand, no consensus is available from randomized trials regarding the optimal total dose and daily fractionation in the different disease stages, although early-stage disease often receives lower doses, while larger and more advanced tumors are treated with higher doses [
1,
2,
3,
4,
5,
6,
7,
8].
Data from randomized trials showed that CRT with relatively low total radiation doses (30–50 Gy) and the addition of MMC to 5FU resulted in high LC rates in small tumors (<4 cm) [
1,
2]. An LC rate ranging between 45–55% has been reported with a modest dose of 45 Gy in 25 fractions and the boost dose modulated according to treatment response [
1,
2,
3,
4]. In the RTOG 8704, 9811 phase III trials (median RT dose of 48 Gy, without planned gap) in patients with T3, T4, node-positive disease and residual tumor, an additional boost of 9 to 14 Gy (2.0 Gy/daily, total dose of 54 to 59 Gy) was delivered to the primary tumor/nodal mass [
4,
6]. Prophylactic irradiation of inguinal lymph nodes up to a dose of 45 Gy increased LC and is recommended, but it remains unclear whether boosting the radiation dose to >50 Gy in patients with good response will improve the results [
4,
6]. Higher doses above 56 Gy may provide better local control, but they could be associated with an increased toxicity [
26], and doses >59 Gy were not shown to provide additional benefits [
27].
In our study, patients were treated with a median total dose of 55 Gy (range: 45–75). SIB was delivered in 568 patients (57.5 %), and an additional sequential EBRT or BRT boost was administered in 167 patients (16.9%). Concomitant chemotherapy was administered in all patients with MMC plus 5-FU or the capecitabine regimen in 779 patients (78.92%).
The treatment was well tolerated, reporting grade 3 acute skin, GI and urogenital toxicity in 251 patients (25.4%), 61 patients (6.2%) and 5 patients (0.5%), respectively. In RTOG 0529, aiming to evaluate the tolerability of a dose-painted IMRT treatment, acute ≥G3 skin and gastrointestinal toxicity rates were 23% and 21%, respectively [
15]. Overall, our analysis showed grade 3–4 acute toxicity in 32.7% of patients, related mainly to local skin toxicity (25.6%). Grade 3–4 acute hematologic toxicity in our study was also limited (8.7%), despite the fact that selective approaches to spare the bone marrow were not routinely used in some institutions [
28].
The good toxicity profile seems noteworthy considering the median OTT of 45 days (range: 25–115), with a treatment interruption of >5 days for toxicity in 186 patients (18.8%). Nine hundred twenty patients (93.2%) completed the planned RT treatment with a high compliance to treatment. These data confirmed the treatment tolerability with a low rate of interruptions with IMRT techniques, even when SIB is delivered with consequent reduced OTT.
The SIB strategy, allowing for both gross tumor volume and elective volumes to receive different total doses in the same number of fractions, could require that different elective nodal volumes are treated with a varying dose per fraction [
14,
15]. Historically, anal cancer has been treated with doses of 1.8 Gy per fraction, using a shrinking field technique over the course of treatment. Using IMRT with SIB to treat different target volumes with different daily doses often results in some areas receiving less than conventional fractional doses of radiation (i.e., <1.80 Gy). In the phase II RTOG 0529 study [
15], some patients received, to some elective nodal volumes, a fractional dose of 1.5 Gy without loss of effectiveness in LC and disease-free survival rates compared to RTOG 9811 [
6]. The median OTT in the study was 43 days compared with 49 days in the RTOG 9811.
In AC patients, the reduction of OTT could lead to a clinical benefit, whereas a longer OTT could be detrimental to the likelihood of receiving a colostomy and LC [
10,
11] due to an accelerated repopulation that could occur after irradiation and may lead to a loss in tumor control [
29]. Data pooling from the RTOG-8704 and the RTOG-9811 studies showed that OTT was positively associated with time to colostomy failure but not with OS or CFS [
10]. Although our analysis did not show a significant correlation between OTT prolongation and CR rate, a detrimental trend effect was observed between OTT > 45 days and PFS (
p = 0.050) and was significantly associated with a lower EFS (
p = 0.030), confirming the benefit of “gap” avoidance techniques, such as IMRT-SIB [
11].
Recently, according to the UK IMRT guidance [
30], a consecutive cohort of 385 patients was retrospectively analyzed. All treatments were delivered by SIB in 28 fractions, with T1-2 N0 receiving 50.4 Gy to the gross primary tumor and T1-2 N+ or T3-4 N-any receiving 53.2 Gy. Involved nodes received 50.4 Gy if 3 cm large; uninvolved pelvic nodes (PTV-elective), including mesorectal, obturator, external and internal iliac, inguinal and presacral, received 40 Gy since March 2014 (dose per fraction: 1.4 Gy). As reported by the authors, this was a biologically equivalent dose to 30.6 Gy in 17 fractions delivered in the ACT2 trial, using an alpha–beta ratio of 8 and a loss of 0.7 Gy per day after 20 fractions. Only two isolated lymph node relapses have been reported with this low dose per fraction approach on elective volume [
31]; then, the efficacy of doses less than 1.80 Gy per day are assumed.
Parallel to these studies, in our cohort the elective low-risk PTV received less than 1.80 Gy per day in more than half of the patients. No statistically significant correlation was found between boost modality (SIB), total RT dose (grouping patients receiving less than, equal to and more than 54 and 55 Gy) or dose per fraction (grouping patients receiving hazard ratio (HR) PTV less than, equal to and more than 2 Gy/fraction, and LR PTV less than, equal to and more than 1.8–2 Gy/fraction) and clinical response and outcomes.
Even though a certain heterogeneity in the dose volume prescription of elective PTVs has been reported in our study, this factor did not affect treatment response, and the several RT schedules used seem to be equivalent in terms of clinical outcomes. Based on these results, although recommendations about optimal total dose and fractionation cannot be provided, our study confirmed that low fractional doses may be appropriate for clinically negative areas when using IMRT for AC with concurrent chemotherapy. The ongoing PLATO integrated protocol looking at dose escalation in LAD and dose de-escalation in early small-node-negative disease is currently set up in the UK, and in the future will inform dose fraction optimization for AC. Finally, we are currently performing a national survey to explore the gray areas in the pattern of care of AC and to define a clinical practice consensus, especially in terms of treatment doses and volumes.
Data on the HPV status of more than half of our patients were missing, which is probably related to the retrospective nature of the study, and this could explain the positive/negative HPV-rate in our analysis. Indeed, AC is reported to be associated with HPV infection in 70–90% of cases, with HPV16 as the most common sub-type. This could also have had an impact on the non-statistically significant correlation with clinical outcomes in the two subgroups.
Histological grade 3 and lymph node involvement (with any T) were associated with a lower probability of CR (OR 0.11, 95% CI 0.03–0.52,
p = 0.030 and OR 0.45, 95% CI 0.25–0.76,
p = 0.004, respectively) and lower LC (
p < 0.001 and
p = 0.025, respectively). Lymph node involvement remained negatively associated with all clinical outcomes (LC, CFS, OS, PFS and EFS) also in the multivariate analysis. Advanced stage was confirmed to be a negative prognostic factor for CFS, OS, PFS and EFS. Furthermore, as reported in other studies, nodal involvement and advanced stage results were significant predictors for LC, OS and PFS [
32]. Moreover, disease extension results, probably due to large volumes treated at high doses, were associated with higher risk of toxicity.