summary of changes
untitled h2

ANCHOR Study: Anal Cancer Prevention Study Anal Cancer/HSIL Outcomes Research Study Version 15.0 NCI Protocol \#: AMC-A01 Local Protocol \#: ANCHOR NCI Version Date: 22JAN2021 Protocol Date: 22JAN2021
summary of changes
i. comments requiring a response

| disapproval letter for protocol version 14:   |                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                      |                                                                             |
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| #                                             | Section                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                              | Comments                                                                    |
| 1.                                            | 8.5.11,                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                              | Response to comment 2 is unclear: the protocol should make it clear whether |
| 8.6.13                                        | missed follow-up evaluation visits are considered a protocol violation.  Please note: CTEP does not approve open-ended missed follow up and  endpoint re-evaluation visits that can affect the primary endpoint analysis.  Guidelines and time limits must be made clear, including when to consider  successive missed visits a protocol deviation.  It would appear that the investigators wish to follow patients regardless of  potential impact on the primary endpoint. If this is the case, then a definition  of when a patient can no longer viably contribute to the primary endpoint must  be included, along with a sensitivity and feasibility analysis for when the  protocol itself can no longer accomplish it's primary goal.  PI Response: There are no stated exemptions from protocol deviations  from missed 6-month visits. We stated in our prior response that these are  unavoidable deviations. If a site performs a telehealth visit because the 6- month visit cannot be performed, it will still require reporting as a missed  visit and is still considered a protocol deviation. This has been  communicated previously to sites per CTEP guidance regarding  management of the effects COVID-19 on research visits. At CTEP's  request, this is now repeated in the protocol. Participants are evaluable for the primary endpoint until they develop a  confirmed anal cancer diagnosis or expire, or withdraw informed consent  for participation. This is described in section 4.5 and the investigators find  no other reason to omit a participant from the primary analysis (ITT). As  stated in the prior response, telehealth visits also do not supplant in-office  visits in determining person-years of follow-up completed. The approach |                                                                             |  
| #                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                        | Section   | Comments   |
|--------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|-----------|------------|
| to determining the amount of follow-up for all cases involves calculating  the duration between the latest date HRA was performed and the  randomization date, and to compare visit completion rates to the expected  visit completion rates based on the participant's visit schedule. The DSMB  reviews this information during its regular reviews of the protocol, in  conjunction with the interim analyses and the dropout rate. While visits  have been missed due to necessary COVID-19 -related restrictions, the  protocol is not at risk of failing to accomplish its primary analysis (last  reviewed by the DSMB November 2020, and to be reviewed at least  annually for the remainder of trial duration). |           |            |  
II. Recommendations from the 22 December 2020 disapproval letter for protocol  
version 14:  
| version 14:   |         |                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                         |
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| #             | Section | Comments                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                |
| 2.            | 8.1.12  | The protocol indicates that collection tubes containing oropharyngeal swabs  from ARAS-CoV-2 testing must be stored at -70C or colder until shipment to  the biorepository (e.g., see page 89). The investigators should confirm that all  sites that will participate in the ancillary SARS-CoV-2 study have the facilities  to store the collection tubes at this temperature. PI Response: The study team qualified the freezers available at each site  before site initiation. If the clinical site does not have a -70C (or colder) refrigerator, the manual of procedures specifies appropriate alternate  procedures (shipping specimens on the same day as collection, or, if  collected on a Friday or before a holiday, storing specimens frozen at -20C  and then shipping on dry ice on the next available business day). The study  specimens may also be stored at -20C and the protocol has been revised  accordingly. This level of detail for these special cases is not required for the  protocol document and is appropriately managed in a manual of  procedures. |
| 3.            | 10.6    | Under Aim 1 in section 10.6 the protocol states: "With a sample size of 400 for  each swab type, the 95% confidence interval for the SARS-CoV-2 prevalence  will be no greater than +/- 0.3%." The 0.3% figure appears incorrect. Under the  worst-case scenario of a prevalence of 0.5 (50%), the 95% CI width would be  5.0%. Possibly the investigators are computing the width under the hypothesized  prevalence of 10%, but in that case the width would be 3.0% and not 0.03%.  Assumptions should be clarified, and corrections made as needed.  PI Response: A key sentence was omitted in this section. We postulated that  the prevalence of SARS-CoV-2 would be 10%; thus, the width of the 95%  confidence interval would be ± 3.0%.                                                                                                                                                                                                                                                                                                                                       |  
III. Comments Requiring a Response - Major issues from the 24 November 2020  
disapproval letter for protocol version 13:  
| disapproval letter for protocol version 13:   |                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                            |                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                |
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| #                                             | Section                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                    | Comments                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                       |
| 1.                                            | 2.3.11                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                     | This section states, "These results provide evidence of A-HRSI clinical  responsiveness that will be further explored as part of the ANCHOR trial  through refined assessment windows". Please clarify what additional  psychometric work on the tool is needed.  PI Response: Please refer to the subsequent sentences that describe how  the windows for these evaluations were refined for the QoL objective. This  sentence introduces this subsequent description for the QoL objective.  Minor phrasing changes were applied so that the reader appropriately  identifies the context of this paragraph. |
| 2.                                            | 8.5.11,                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                    | Amended sections 8.5.11 and 8.6.13 allow replacing follow-up evaluation                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                        |
| 8.6.13                                        | visits with a virtual visit. This may compromise evaluation of the primary  endpoint and hurt study ability to address its primary goal. Every effort should  be made to maintain the follow up visit schedule (missed visits should be  rescheduled, and if a visit cannot be rescheduled it should be considered  protocol non-compliance).  PI Response: As stated in our prior response, the study team's intent is  that every effort must be made to maintain the follow-up visit scheduled.  No waiver for the 6-month visit is provided with these changes. Missed  visits due to COVID-19 may be unavoidable deviations due to local site  closures or restrictions. The intent of this provision is to provide a  provision for telehealth when in-person visits are not possible or the  participant is unwilling to attend a visit, for the purposes of collecting  follow up information for safety, study arm adherence, supporting  participant retention, and determining whether the participant has any  anal symptoms that may warrant an immediate visit for the participant's  care. Clinic staff report that this process supports retention by  maintaining communication with the participant when the 6-month visit  cannot be scheduled. Sites are still instructed to conduct a makeup 6- month visit once this is feasible again. This statement has been added to  the protocol to emphasize this intent. The study team believes this process minimizes the effect of COVID-19 on  the primary endpoint by allowing clinicians to remain in contact with  participants when visits are not feasible and to identify participants who  require immediate care due to concern for potential cancer, on the basis  of reported symptoms. The study team recognizes this information cannot  supplant evaluation for the primary endpoint by HRA with biopsy, but in  the presence of pandemic-related restrictions preventing visit conduct, this  is the best available approach to weigh when the potential benefit of  conducting a visit or providing care outweighs the risk of SARS-CoV-2 |                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                |  
| #                                                                                                                                                                                                                                                                                                                                            | Section   | Comments                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                 |
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| transmission in the clinic. The information from these follow-up calls are  collected in a separate case report form in the database that will not affect  the follow-up calculation for the primary analysis; this will be based on the  time elapsed between the participant's randomization date and the date of  the last HRA performed. |           |                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                          |
| 3.                                                                                                                                                                                                                                                                                                                                           | 10.1.2    | It is not clear why this is an secondary objective: To assess the responsiveness  (sensitivity to change) and clinical significance of the A-HRSI subscales by  comparing change scores within groups of participants as defined by  participant responses to the PGIC item (n=500). Hasn't this work been  completed in February with a sample size of 100 patients?  PI Response: This was a copy/paste error that has been deleted. See section  10.1.4 for the QoL objectives, which are also stated in section 1.5. |
| 4.                                                                                                                                                                                                                                                                                                                                           | 10.5      | There is no explanation of how missing data will be handled.  PI Response: The sample sizes for the QoL substudy and QoL objective  account for rates of non-response and dropout, based on the substudy. In  addition to the specified analyses that assume data are missing at random,  multiple imputation methods will be applied as a sensitivity analysis to  assess the impact of missing data that may not be missing at random.                                                                                 |
summary of changes
iv. comments requiring a response

| #   | Section   | Comments                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                              |
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| 5.  | 2.3.11    | Aims of the A-HRSI substudy were to validate the HR-QoL questionnaire-final  data collection was completed in February 2020 and final results are still being  analyzed. Based on the interim results, the questionnaire is now being  implemented in the ANCHOR study. However, the results of the psychometric  work are not reported in the protocol. Please provide the results, scoring,  responsiveness over time and the change in score that is considered clinically  meaningful. There is no information regarding clinical significance of  calculated non-normalized ES for the A-HRSI subscales.  PI Response: Results from the A-HRSI substudy have been added to the  background section (section 2.3.11). In section 10.5, we comment on the  minimum differences we can detect based on our sample size calculations for the final implementation of A-HRSI as compared to those seen in the  responsiveness substudy when comparing those with worse ECOG PS  scores versus those with no change. The A-HRSI substudy provided a  preliminary indication of change that would be considered clinically  meaningful. |  
| #                                                                                                                                                                                                                                                                                                                                               | Section                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                         | Comments                                                                                                                                                                                                                                                                                                                                                    |       |    |            |     |        |               |    |       |    |
|-------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|-----------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|-------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|-------|----|------------|-----|--------|---------------|----|-------|----|
| 6.                                                                                                                                                                                                                                                                                                                                              | 4.1                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                             | Section 4.1 states patients will undergo HRA every 6 months, and biopsy of  HSIL every 12 months. New sections 8.5.1 and 8.6.13 provide for visit deferrals  with the possibility of telemedicine consultation instead. There seems to me  limit on length of deferrals on these follow-up exams, or provisions for  scheduling a deferred follow-up visit. |       |    |            |     |        |               |    |       |    |
| 8.5.10,  8.6.13                                                                                                                                                                                                                                                                                                                                 | Please reconcile section 4.1 with the appropriate sections in 8, and provide  more direction for how much deferral can occur before requiring patients to  come off study.  PI Response: Per the guidance of the ANCHOR DSMB during its second  visit in December 2015, participants should not be taken off study unless  they expire or their disease satisfies the primary endpoint (progression to  cancer). As this is a time to event study, if a participant remains willing  and able to return to the trial, a visit should be scheduled to perform HRA  with biopsy regardless of whether prior visits were missed or performed  as a telemedicine follow-up visit. Telemedicine visits during the COVID  pandemic are a means to continue to collect data on participant safety and  study arm compliance if an in-person visit is not feasible due to COVIDrelated restrictions only.                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                 |                                                                                                                                                                                                                                                                                                                                                             |       |    |            |     |        |               |    |       |    |
| 7.                                                                                                                                                                                                                                                                                                                                              | 8.2, 8.7                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                        | There is not justification provided for the 9 timepoints of measurement.                                                                                                                                                                                                                                                                                    |       |    |            |     |        |               |    |       |    |
| 2.3.11                                                                                                                                                                                                                                                                                                                                          | PI Response: As discussed during a teleconference with representatives  from CTEP on 25AUG2020, time 2 (T2) will be revised to occur 2-7 days  after randomization, to correspond with the period during which most  participants are symptomatic after the initial treatment or monitoring arm  HRA, and time 3 (T3) has been revised to occur at 4 weeks after  randomization, at which point the investigators would not expect  participants to be symptomatic. Subsequent time points 4-8 are now  scheduled to occur around annual visits through the fifth year of study  participation, to evaluate any long-term psychological effects of each study  arm's strategy. Time 1 (baseline) remains within 2 weeks before  randomization. Time points 4-8 will be used in an exploratory analysis in  order to characterize prospective changes in HR-QoL. |                                                                                                                                                                                                                                                                                                                                                             |       |    |            |     |        |               |    |       |    |
| 8.                                                                                                                                                                                                                                                                                                                                              | 8.4.7                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                           | The                                                                                                                                                                                                                                                                                                                                                         | value | of | completing | the | HR-QoL | questionnaire | 2  | weeks | post    |
| randomization is unclear. Is there an expected change from the baseline  measurement in such a short period of time?  PI Response: Please see our responses to items #3 and 6 for the rationale for time point 2 and its role as the expected timeframe in which physical  effects of the monitoring and treatment strategies will be measured. |                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                 |                                                                                                                                                                                                                                                                                                                                                             |       |    |            |     |        |               |    |       |    |
| 9.                                                                                                                                                                                                                                                                                                                                              | 8.6.10.1                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                        | Please explain the new provision to allow treatment of HSIL if it was biopsied  6 months prior.                                                                                                                                                                                                                                                             |       |    |            |     |        |               |    |       |    |  
| #   | Section   | Comments PI Response: This provision permits empirical treatment of clinically  suspected HSIL lesions in the treatment only arm only if the biopsies are  collected at the same visit as they are treated and not the prior six month  visit. The new provision allows clinicians to treat these lesions to avoid  having to biopsy the lesion, wait for the result and bring the participant  back at a separate visit for treatment, as long as their suspicion of cancer  is low. To confirm they that did not unintentionally treat a cancer, we do  however require that they biopsy the lesion before treating it.                                                                                                                                                                                                                                                                                                                                                                                                                         |
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| 10. | 10.5.2    | There is no stated hypothesis for the HR-QoL sub-study.  PI Response: The primary endpoint for the QoL objective is changes in  physical symptoms and impacts from T2 to T3, adjusting for T1. It is  anticipated that participants assigned to the treatment arm will experience  significant reduction in physical symptoms and impacts from T2 to T3  versus those in the active monitoring arm. The secondary endpoint for the  QoL objective is changes in psychological symptoms from T3 to T4,  adjusting for T1. There is uncertainty of the timing of psychological  symptoms related to lack of treatment among those in the active  monitoring arm, but it is anticipated that participants assigned to the  active monitoring arm will experience significant increases in  psychological symptoms from T3 to T4, a later time point compared with  those in the treatment arm. The exploratory endpoint is HR-QoL changes  in physical symptoms/impacts and psychological symptoms from T4  through the subsequent T5-T8 follow-ups. |
| 11. | 10.5.2    | The HR-QoL study endpoint is not clear. The investigators state they will  perform mixed model repeated measures ANOVA with change at baseline  calculated at each follow up visit for each subscale. Do the investigators intend  to do a longitudinal analysis? What comparison and timepoint(s) are of interest?  PI Response: Please see our response to item #6. The analyses for these  endpoints are detailed in section 10.5.2.                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                           |
| 12. | 10.5.2    | It is not appropriate to use 5% two-sided alpha per sub-scale, because of  multiplicity of analyses. The overall type one error should be controlled for at  5% level. PI Response: For the primary endpoint, we are testing two subscales and  anticipate that participants assigned to the treatment arm will experience  significant reduction in physical symptoms and impacts from T2 to T3 versus  those in the active monitoring arm. Therefore, we will conduct two tests at the  one-sided alpha level of 0.025, based on applying a Bonferroni correction to  maintain an overall one-sided alpha level of 0.05.                                                                                                                                                                                                                                                                                                                                                                                                                        |
| 13. | 10.5.2    | For responsiveness, the investigators consider the possibility of a scenario  leading to type one error +0.31. If this scenario is realistic, analysis does not                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                   |
section comments
iv. comments requiring a response

| make sense. PI Response: This language was not a part of the current amendment for  the main QoL objective but was related to the responsiveness substudy. It  was intended to show increased Type I error if effect sizes were smaller  than what the substudy was designed around.   |
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| V.   | Scientific and Substantive Changes                                                                  |                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                   |
|------|-----------------------------------------------------------------------------------------------------|---------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|
| #    | Section                                                                                             | Description of Change                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                             |
| 14.  | Synopsis 1.6 2.6 3.5 3.6 3.7 8.1.12 8.2 8.5 10.1 10.6 13.0 Appendix I                               | The protocol has been revised throughout to add a supplemental study  to describe detection of SARS-CoV-2 in anal swab samples from  PLWH being screened for the ANCHOR study; examine its  relationship to prevalent anal HPV infection and HSIL in the  screening population; and determine its effect on the natural history of  anal HPV infection and HSIL by examining its relationship to  regression of HSIL and clearance of HPV infection in the subset of  enrolled participants randomized to the active monitoring arm.  Accordingly, our specific aims are outlined in Section 1.6. |
| 15.  | Synopsis Schema 1.5  2.3.11 2.4 2.4.1 2.4.2 3.3 3.4 3.6.1 3.7 8.3 8.4 8.5 8.6 8.7 8.8 8.9 10.1 10.2 | The protocol was revised to report completion of the A-HRSI scale  responsiveness substudy and to implement the A-HRSI instrument in  the protocol as a formal QoL aim. The objectives, eligibility criteria,  study procedures, statistical section, and appendices were revised  accordingly. The QoL aim will also include collection of data using  the Spanish language version of the A-HRSI as validated in the AMCA04 protocol, Development of a Health-Related Symptom Index for  Spanish-Speaking Persons Diagnosed with and either Treated or  Monitored for Anal High-Grade Squamous Intraepithelial Lesions  (HSIL).                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                   |  
| #                                                            | Section                                                                                                                                                                                                                                                            | Description of Change                                                                                                                                                                                                                                          |                 |           |             |        |         |    |       |
|--------------------------------------------------------------|--------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|----------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|-----------------|-----------|-------------|--------|---------|----|-------|
| 10.4                                                         |                                                                                                                                                                                                                                                                    |                                                                                                                                                                                                                                                                |                 |           |             |        |         |    |       |
| Appendix I Appendix X Appendix XI Appendix XII Appendix XIII |                                                                                                                                                                                                                                                                    |                                                                                                                                                                                                                                                                |                 |           |             |        |         |    |       |
| 16.                                                          | 2.3                                                                                                                                                                                                                                                                | A definition for an anal cancer case that qualifies for the primary  endpoint was added to the protocol, based on current National  Comprehensive Cancer Network (NCCN) guidelines for the extent of  disease that is considered anal squamous cell carcinoma. |                 |           |             |        |         |    |       |
| 17.                                                          | 4.3                                                                                                                                                                                                                                                                | The                                                                                                                                                                                                                                                            | exclusion       | criterion | prohibiting | recent | receipt | of | other |
| investigational                                              | drugs                                                                                                                                                                                                                                                              | (excepting                                                                                                                                                                                                                                                     | investigational | ART       | agents,     |        |         |    |       |
| treatments for Hepatitis C, and treatments/vaccinations for SARSCoV-2) has been extended as prohibited therapy during study  participation.                                                              |                                                                                                                                                                                                                                                                    |                                                                                                                                                                                                                                                                |                 |           |             |        |         |    |       |
| 18.                                                          | 6.4                                                                                                                                                                                                                                                                | Data on COVID-19 diagnoses (suspected and confirmed) will now be  collected as routine adverse events, for the purpose of identifying  cases in the future as needed for ancillary research proposals in  development.                                         |                 |           |             |        |         |    |       |
| 19.                                                          | 8.5.11                                                                                                                                                                                                                                                             | Telephone follow-up procedures were added to the protocol for 6-                                                                                                                                                                                               |                 |           |             |        |         |    |       |
| 8.6.13                                                       | month visits unable to be conducted due to COVID-19, to formalize  the study's procedures for maintaining contact with participants and  evaluating patient safety and adherence to the assigned study arm to  the extent feasible during the SARS-CoV-2 pandemic. |                                                                                                                                                                                                                                                                |                 |           |             |        |         |    |       |
| 20.                                                          | Appendix III                                                                                                                                                                                                                                                       | Guidance regarding insufficient or inconclusive cytology at screening  was added, requiring repeat testing if it occurs. Valid results must be  available prior to randomization (may occur up to 12 weeks prior to  randomization).                           |                 |           |             |        |         |    |       |
section comments
vi. administrative and editorial changes

| #   | Section   | Description of Change                                                                                          |
|-----|-----------|----------------------------------------------------------------------------------------------------------------|
| 21. | Global    | The protocol version and version date were updated to version 15.0,  dated 22JAN2021.                          |
| 22. | Global    | Spelling, grammar, and editorial changes (abbreviations, company  names) were applied throughout the document. |
| 23. | Global    | The manufacturer name for topical medications was updated from  Valeant Pharmaceuticals to Bausch Health.      |  
| #                    | Section                                                                                                    | Description of Change                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                             |                          |           |
|----------------------|------------------------------------------------------------------------------------------------------------|-----------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|--------------------------|-----------|
| 24.                  | List of Tables                                                                                             | Lists of tables and figures were added to the protocol, and each table                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                            |                          |           |
| List of Figures      | and figure renumbered accordingly.                                                                         |                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                   |                          |           |
| 25.                  | 2.1.1                                                                                                      | The background section                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                            | was updated with current | anal HSIL |
| incidence estimates. |                                                                                                            |                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                   |                          |           |
| 26.                  | 8.5.9.1                                                                                                    | Statements regarding empiric treatment of HSIL in the treatment arm                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                               |                          |           |
| 8.6.10.1             | following biopsy at a given visit were repeated in the evaluations  section at visits after randomization. |                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                   |                          |           |
| 27.                  | Appendix VI                                                                                                | The AMC Data and Safety Monitoring Plan, which applies to all AMC  protocols, was updated from version 6.0 to version 9.0. Key revisions  include: the addition of an introduction to address the variety of  systems the AMC uses for individual trials; changes to the data entry  systems used by domestic AMC trials (OPEN/Rave) activated  September 1, 2020 and later; participation with a single IRB for new  domestic AMC protocols; updated procedures for data reporting,  determination of requirements for DSMB review, and safety  review/pharmacovigilance; and administrative changes (updates to  document organization, external links, and group terminology). |                          |           |  
ANCHOR Study: Anal Cancer/HSIL Outcomes Research Study  
  
(AMC Protocol \#A01)
A Collaboration of the AIDS Malignancy Consortium  
| Human Papillomavirus Working Group                                     |                                                                    |
|------------------------------------------------------------------------|--------------------------------------------------------------------|
| and the University of California, San Francisco                        |                                                                    |
| Sponsored by:                                                          | National Cancer Institute Office of HIV and AIDS Malignancy (OHAM) |
| NCT Registration Number:                                               | NCT02135419                                                        |
| Regulatory Status:                                                     | IND exempt drugs IDE exempt study devices                          |
| Commercially Available                                                 | 5-Fluorouracil (5-FU) (NSC 19893)                                  |
| Agents:                                                                | Topical Imiquimod Cream (5%) (NSC 741062)                          |
| Protocol Chair:                                                        | Joel Palefsky, MD, CM, F.R.C.P.(C)                                 |
| Protocol Co-Chair:                                                     | J. Michael Berry, MD                                               |
| AMC-A01 Version 15.0, 22 January 2021 NCI Version Date 22 January 2021 |                                                                    |
protocol signature page
vi. administrative and editorial changes

I, , Principal Investigator at site _______________, agree to conduct and follow this protocol: **ANCHOR Study (AMC-A01, Version 15.0, 22JAN2021),** as written according to NCI, OHRP, and FDA guidelines, and group policies. I understand that no deviations from the protocol eligibility criteria or waivers for protocol deviations will be permitted. PI Signature Date (mm/dd/yyyy)
protocol signature page
table of contents

| SUMMARY OF CHANGES .                                                   |                                                                                                                                                    |
|------------------------------------------------------------------------|----------------------------------------------------------------------------------------------------------------------------------------------------|
| Comments Requiring a Response - Major issues from the 22 December 2020 |                                                                                                                                                    |
| I.                                                                     | disapproval letter for protocol version 14:                                                                                                        |
| II.                                                                    | Recommendations from the 22 December 2020 disapproval letter for protocol                                                                          |
| version 14:                                                            |                                                                                                                                                    |
| II.                                                                    | Comments Requiring a Response - Major issues from the 24 November 2020                                                                             |
| disapproval letter for protocol version 13:                            |                                                                                                                                                    |
| IV.                                                                    | Comments Requiring a Response - Major Issues from the 25 June 2020                                                                                 |
| Disapproval Letter for protocol version 12:                            |                                                                                                                                                    |
| V.                                                                     | Scientific and Substantive Changes ....................................................................................................vii         |
| VI.                                                                    | Administrative and Editorial Changes .                                                                                                             |
| PROTOCOL SIGNATURE PAGE                                                |                                                                                                                                                    |
| TABLE OF CONTENTS                                                      |                                                                                                                                                    |
| LIST OF TABLES .                                                       |                                                                                                                                                    |
| LIST OF FIGURES                                                        |                                                                                                                                                    |
| PROTOCOL ROSTER                                                        |                                                                                                                                                    |
| PROTOCOL SYNOPSIS                                                      |                                                                                                                                                    |
| PROTOCOL SCHEMA                                                        |                                                                                                                                                    |
| 1.0                                                                    | OBJECTIVES                                                                                                                                         |
| 1.1                                                                    | Hypothesis                                                                                                                                         |
| 1.2                                                                    | Primary Objective                                                                                                                                  |
| 1.3                                                                    | Secondary Objectives                                                                                                                               |
| Exploratory Objectives .                                               |                                                                                                                                                    |
| 1.4                                                                    | Quality of Life Objectives .....................................................................................................................20 |
| 1.5                                                                    |                                                                                                                                                    |
| 1.6                                                                    | Ancillary Study Objectives .                                                                                                                       |
| 2.0                                                                    | BACKGROUND                                                                                                                                         |
| 2.1                                                                    | Anal Cancer                                                                                                                                        |
| 2.2                                                                    | Study Agents and Treatments                                                                                                                        |
| 2.3                                                                    | Study Design and Rationale                                                                                                                         |
| 2.4                                                                    | Overview of Study Design .                                                                                                                         |
| Correlative Studies                                                    |                                                                                                                                                    |
| 2.5                                                                    |                                                                                                                                                    |
| 2.6                                                                    | Ancillary Study: SARS-CoV-2 and Anal HPV Infection                                                                                                 |
| 3.0                                                                    | PATIENT SELECTION                                                                                                                                  |  
| 3.1                                              | Eligibility Criteria 60                                                                                                                                                                                                                                                                                                            |
|--------------------------------------------------|------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|
| 3.2                                              | Exclusion Criteria 62                                                                                                                                                                                                                                                                                                              |
| 3.3                                              | Inclusion Criteria for the HRQoL Substudy 62                                                                                                                                                                                                                                                                                       |
| 3.4                                              | Inclusion Criteria for the QoL Objective 63                                                                                                                                                                                                                                                                                        |
| 3.5                                              | Inclusion Criteria for the SARS-CoV-2 Ancillary Study 63                                                                                                                                                                                                                                                                           |
| 3.6                                              | Number of Participants to be Enrolled 63                                                                                                                                                                                                                                                                                           |
| 3.7                                              | Participant Enrollment Procedures 63                                                                                                                                                                                                                                                                                               |
| 4.0                                              | TREATMENT PLAN 65 4.1 Group 1: Active Monitoring Arm 65 4.2 Group 2: Treatment Arm 65 4.3 General Concomitant Medication and Supportive Care Guidelines 71 4.4 Concurrent HPV-related Disease Treatment 72 4.5 Duration of Follow-Up 73 4.6 Criteria for Removal from the Study (Drop-Out) 74                                      |
| 5.0                                              | DOSING DELAYS/DOSE MODIFICATIONS 75 5.1 Dose Modifications Guidelines 75                                                                                                                                                                                                                                                           |
| 6.0                                              | ADVERSE EVENTS: LIST AND REPORTING REQUIREMENTS 76 6.1 Potential Risks Lists 76 6.2 Adverse Event Characteristics and Definitions 77 6.3 Expedited Adverse Event Reporting 79 6.4 Routine Adverse Event Reporting 81 6.5 Secondary Malignancy 82 6.6 Second Malignancy 82                                                          |
| 7.0                                              | PHARMACEUTICAL AND STUDY DEVICE INFORMATION 83 7.1 5-Fluorouracil 5% Cream 83 7.2 Topical Imiquimod Cream (5%) 84 7.3 Study Devices 85 7.4 Drug Supply and Accountability 85                                                                                                                                                       |
| 8.0                                              | CLINICAL AND LABORATORY EVALUATIONS 87 8.1 Special Instructions and Definitions of Evaluations 87 8.2 Screening Evaluations (Visit 0) 90 8.3 Baseline Visit with Randomization Evaluations (Visit 1) 91 8.4 Evaluations between Six Month Visits (Interim Visits) 93 8.5 Evaluations at Visit 2 (6 Months) for All Participants 93 |
| ANCHOR Protocol (AMC-A01 Version 15.0) 22JAN2021 | 4                                                                                                                                                                                                                                                                                                                                  |  
4  
| 8.6                                                                                                                                                                                                                                                                                                                   | Evaluations at Follow-Up (Visits 3-10+), Every 6 Months until Study  Completion 94                                                                                                                                                                                                                                                                         |
|-----------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|
| 8.7                                                                                                                                                                                                                                                                                                                   | Final Evaluations, Off Study 96                                                                                                                                                                                                                                                                                                                            |
| 8.8                                                                                                                                                                                                                                                                                                                   | Early Study Discontinuation Evaluations 97                                                                                                                                                                                                                                                                                                                 |
| 8.9                                                                                                                                                                                                                                                                                                                   | QoL Objective 98                                                                                                                                                                                                                                                                                                                                           |
| 9.0                                                                                                                                                                                                                                                                                                                   | BIOMARKER STUDIES 99 9.1 Viral Factors in HSIL Progression to Cancer 99 9.2 Identification of Host Factors in HSIL Progression to Cancer 101 9.3 Viral and Host Biomarker Predictors of Risk of Progression from HSIL to  Cancer 104 9.4 Medical History and Behavioral Risk Factors for Progression from HSIL to  Cancer among Those Who Are Enrolled 110 |
| 10.0                                                                                                                                                                                                                                                                                                                  | STATISTICAL CONSIDERATIONS 112 10.1 Study Design/Endpoints 112 10.2 Sample Size/Accrual Rate 112 10.3 Stratification Factors 115 10.4 Statistical Analysis 115 10.5 Quality of Life Analyses 116 10.6 SARS-CoV-2 Ancillary Study Analyses 118 10.7 Data and Safety Monitoring 119                                                                          |
| 11.0                                                                                                                                                                                                                                                                                                                  | ROLE OF DATA MANAGEMENT 122 11.1 CRF Instructions 122 11.2 Data Quality 122 11.3 Data Monitoring 122                                                                                                                                                                                                                                                       |
| 12.0                                                                                                                                                                                                                                                                                                                  | ETHICAL AND REGULATORY CONSIDERATIONS 123 12.1 IRB Approval and Informed Consent 123 12.2 Changes to the Protocol 123 12.3 Women and Minorities 123                                                                                                                                                                                                        |
| 13.0                                                                                                                                                                                                                                                                                                                  | REFERENCES 125                                                                                                                                                                                                                                                                                                                                             |
| APPENDIX I: SCHEDULE OF EVALUATIONS 137 Participants Randomized to Active Monitoring Arm 137 Participants Randomized to Treatment Arm, Treatment with Imiquimod Or 5- Fluorouracil Cream 139 Participants Randomized to Treatment Arm, Treatment with Infrared Coagulation,  Hyfrecation/Electrocautery, or Laser 141 |                                                                                                                                                                                                                                                                                                                                                            |  
| Participants Randomized to Treatment Arm, Surgery 143 Health-Related Quality of Life (QoL) Objective Schedule of Evaluations 145                                                                                                                             |
|--------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|
| APPENDIX II: PERFORMANCE STATUS SCALES 146 APPENDIX III: ANAL CYTOLOGY AND HPV SAMPLING 147 APPENDIX IV: HIGH RESOLUTION ANOSCOPY (HRA) AND ANAL BIOPSIES 148 APPENDIX V: RISK FACTOR QUESTIONNAIRE 151 APPENDIX VI: AMC DATA AND SAFETY MONITORING PLAN 156 |  
APPENDIX VII: PARTICIPANT INSTRUCTIONS FOR APPLYING IMIQUIMOD 5% CREAM 161 APPENDIX VIII: PARTICIPANT INSTRUCTIONS FOR APPLYING EFUDEX (5- FLUOROURACIL) 5% CREAM .................................................................................................. 162 APPENDIX IX: REGULATORY STATUS OF INVESTIGATIONAL AGENTS ................... 163 APPENDIX X: ANCHOR HEALTH-RELATED SYMPTOM INDEX..................................... 141 APPENDIX XI: ANCHOR HEALTH-RELATED SYMPTOM INDEX - SPANISH LANGUAGE VERSION ................................................................................................................. 143 APPENDIX XII: PATIENT VERSION OF THE EASTERN COOPERATIVE GROUP PERFORMANCE STATUS............................................................................................................ 145 APPENDIX XIII: PARTICIPANT GLOBAL IMPRESSION OF CHANGE (PGIC) ............. 146
list of tables
table of contents

Table 2-A: Descriptive statistics for change in subscales (T3 minus T2) according to change in PGIC and ECOG PS, mean (SD) ...................................................................................................... 43 Table 2-B: Responsiveness comparison of changes (T3 minus T2), mean difference (95% confidence interval) ............................................................................................................................ 43  
| Table 2-C: A-HRSI completion time points for QoL objective 48 Table 5-A:1. Dose modifications 75   |
|----------------------------------------------------------------------------------------------------|  
Table 6-A: Mapping adverse event attribution assignments to causal relationship designations .. ..................................................................................................................................... 79 Table 10-A: Power associated with varying drop-out rates and drop-in rates from the active monitoring arm with fixed sample size (N=5058), two-sided significance level of 0.05, incidence rate for the active monitoring arm (200/100,000) and the treatment arm (50/100,000) ............ 113 Table 10-B: Sample size = 5058, two-sided significance level = 0.05, 3 year accrual, 5 year follow-up ................................................................................................................................... 114 Table 10-C: Power associated with varying drop-out rates and drop-in rates from the active monitoring arm with fixed sample size (N=5058), two-sided significance level of 0.05, incidence rate for the active monitoring arm (200/100,000) and the treatment arm (50/100,000) ............ 115 Table 10-D: Smallest differences that can be detected for comparing change for specified time points according to treatment group for various scenarios based on pilot data with 90% power using a one-sided alpha of 0.025 ..................................................................................................... 117 Table 10-E: Statistical test boundaries and associated operating characteristics. ..................... 120 Table 12-A: Accrual targets ............................................................................................................ 124
list of figures
table of contents

Figure 2-D Paired Lesion Comparisons ........................................................................................... 51 Figure 4-A: Participant initiates imiquimod/5-FU at visit 1 (randomization visit) ..................... 70 Figure 4-B: IRC, hyfrecation, or electrocautery performed at visit 1 (randomization visit)...... 70 Figure 6-B: Timeframes and Exceptions for Expedited SAE Reporting ...................................... 80
protocol roster
anchor anal cancer prevention study

Protocol Chair: **Protocol Statistician:** Joel Palefsky, MD, CM, F.R.C.P.(C) 513 Parnassus Ave Box 0654 University of California, San Francisco San Francisco, CA 94143 Tel: 415-476-1574 Fax: 415-476-9364 Email: jpalefsky@medicine.ucsf.edu  
Protocol Co-Chair: **Data Management/Operations:** J. Michael Berry, MD ANCHOR Data Management Center 1600 Divisadero Street Room A740 The Emmes Company, LLC University of California, San Francisco 401 N. Washington Street, Suite 700 San Francisco, CA 94115 Rockville, MD 20850 Tel: 415-353-7443 Tel: 301-251-1161 Fax: 415-353-7464 Fax: 301-830-6799 Email: JMichael.Berry@ucsf.edu Email: anchordmc@emmes.com  
National Coordinator: **Arizona Regional ACSR Biorepository:** Abigail Arons, MPH Monika Schmelz, PhD  
University of California, San Francisco Assistant Professor of Pathology  
Mission Hall: Global Health & Clinical Sciences Building  
Department of Pathology  
550 16th Street, 3rd Floor, UCSF Box 0886 1501 N. Campbell Ave. Room 5103 San Francisco, CA 94158-2549 P.O. Box 245043  
Tel: (415) 514-8155 Tucson, AZ 85724  
Fax: (415) 476-5348 Tel: 520-626-8864 Email: abigail.arons@ucsf.edu Fax: 520-626-6081  
Email: schmelz@pathology.arizona.edu  
Jeannette Lee, PhD 4301 West Markham, \#781 Ed III, Room 3212 University of Arkansas for Medical Sciences Little Rock, AR 72205 Tel: 501-526-6712 Fax: 501-526-6729 Email: jylee@uams.edu
protocol synopsis
anchor anal cancer prevention study

Title: ANCHOR Anal Cancer Prevention Study Phase of Study: Phase III  
| Participating Institutions:   | This protocol will be open to sites that have been approved for  participation by the ANCHOR Coordinating Committee and have  at least 2 practitioners certified in high resolution anoscopy (HRA)  and 1 practitioner certified in HSIL treatment (infrared coagulation  (IRC), hyfrecation/electrocautery, or laser ablation) by the  ANCHOR HRA Training and Certification Committee. At least  fifteen sites will be targeted for participation.                                                                                                                                                                                                                                                                              |
|-------------------------------|-----------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|
| Accrual Target:               | 5,058 participants                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                |
| Population:                   | Men and women age 35 and older with HIV infection and biopsyproven high-grade squamous intraepithelial lesions (HSIL) at  baseline. Eligible participants will have no history of anal cancer,  or treatment or removal of HSIL. Approximately 17,385  participants who provide informed consent for study participation  will be screened to identify 5,058 eligible participants with  previously untreated HSIL.                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                   |
| Regimen:                      | Eligible participants will be randomized to treatment or active  monitoring at baseline. Participants will be followed every six  months for HSIL outcomes for up to five years after the last  participant's date of randomization. Throughout the study, the  incidence of invasive cancer in both arms will be monitored, and  biospecimens and associated participant data will be collected for  correlative science studies. Effects of the treatment and monitoring  strategies on participants' quality of life will be assessed using the  ANCHOR Health Related Symptom Index (A-HRSI) in a subset of  consenting participants (n=500) before randomization and at 7 time  points through 5 years after randomization). |
| Ancillary Studies:            | Anal shedding of SARS-CoV-2: relationship to anal HPV infection  and high-grade squamous intraepithelial lesions in people living  with HIV (Availability of Urgent Competitive Revision and  Administrative Supplements on Coronavirus Disease 2019, PA-18- 59), Section 2.6                                                                                                                                                                                                                                                                                                                                                                                                                                                     |
| Duration:                     | Minimum of 5 years of follow-up for each participant                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                              |  
Primary Objective: To determine whether treating anal high-grade squamous  
intraepithelial lesions (HSIL) is effective in reducing the incidence of anal cancer in HIV-infected men and women.  
Secondary Objectives: - To determine the safety of infrared coagulation (IRC),  
electrocautery, imiquimod, laser and 5- fluorouracil treatments for anal HSIL.  
- To assess the responsiveness (sensitivity to change) and clinical significance of the ANCHOR Health-Related Symptom Index (A-HRSI) subscales by comparing change scores within groups of participants as defined by participant responses to the participant global impression of change (PGIC) item.  
Exploratory Objectives: Collect clinical specimens and data to create a bank of wellannotated specimens that will enable correlative science:
- Identification of viral factors in HSIL progression to cancer;
- Identification of host factors in HSIL progression to cancer;
- Identify host and viral biomarkers of progression from HSIL to cancer;
- Identify medical history and behavioral risk factors for HSIL
progression to cancer.  
Quality of Life Objectives: Primary QOL Objective: To compare arms in terms of changes in physical symptoms and impacts from T2 to T3, adjusting for T1. Secondary QOL Objective: To compare arms in terms of changes in psychological symptoms from T3 to T4, adjusting for T1. Exploratory QOL Objective: To assess of long-term HR-QoL changes in physical symptoms/impacts and psychological symptoms from T4 through the subsequent T5-T8 follow-ups overall and by arm.  
Ancillary Study Objectives:
1. Determine the prevalence of SARS-CoV-2 detection in anal and oropharyngeal swabs among people living with HIV (PLWH) being screened for the ANCHOR study 2. Determine the relationship in the ANCHOR screening population between prevalent anal SARSCoV-2 positivity, anal HPV infection, and anal high-grade squamous intraepithelial lesions (HSIL)  
3. Determine the 6-month incidence of SARS-CoV-2 detection in anal and oropharyngeal swabs among participants with anal HSIL newly enrolled into the ANCHOR study 4. Determine the relationship between prevalent or incident SARS-CoV-2 detection and regression of anal HPV infection or HSIL among participants newly enrolled into the ANCHOR study and who are randomized to the monitoring arm
PROTOCOL SCHEMA  
  
  
  
  
  
  
  
  
ANCHOR protocol modification (v13) to administer  
  
validated A-HRSI in a sample (n=500) of randomized participants at 8 time points: baseline, 2-7 days post randomization, 4 weeks post randomization, and at 12, 24, 36, 48, and 60 months after randomization  
| LIST OF ABBREVIATIONS   |                                                                  |
|-------------------------|------------------------------------------------------------------|
| 5-FU                    | 5-fluorouracil                                                   |
| A-HRSI                  | ANCHOR Health-Related Symptom Index                              |
| ACSR                    | AIDS and Cancer Specimen Resource                                |
| ADL                     | Activities of daily living                                       |
| AE                      | Adverse event                                                    |
| AIDS                    | Acquired immunodeficiency syndrome                               |
| AIN                     | Anal intraepithelial neoplasia                                   |
| AM                      | Active monitoring                                                |
| AMC                     | AIDS Malignancy Consortium                                       |
| AML                     | Acute myelocytic leukemia                                        |
| AMC                     | AIDS Malignancy Consortium                                       |
| ANC                     | Absolute neutrophil count                                        |
| ANCHOR                  | Anal Cancer/HSIL Outcomes Research Study                         |
| APOT                    | Amplification of Papillomavirus Oncogene Transcript              |
| ART                     | Antiretroviral therapy                                           |
| ASCCP                   | American Society for Colposcopy and Cervical Pathology           |
| ASIL                    | Anal squamous intraepithelial lesions                            |
| AWE                     | Acetowhite epithelium                                            |
| BID                     | Bis in die (twice a day)                                         |
| cART                    | Combination antiretroviral therapy                               |
| CBC                     | Complete blood count                                             |
| CDC                     | Centers for Disease Control and Prevention                       |
| CDUS                    | Clinical Data Update System                                      |
| CLIA                    | Clinical Laboratory Improvement Amendments                       |
| CR                      | Complete response                                                |
| CRT                     | Chemoradiation therapy                                           |
| CTCAE                   | Common Terminology Criteria for Adverse Events                   |
| CTEP                    | Cancer Therapy Evaluation Program                                |
| CTEP-AERS               | Cancer Therapy Evaluation Program Adverse Event Reporting System |
| CTMS                    | Clinical Trials Monitoring Service                               |  
| DAB   | Diaminobenzidine                            |
|-------|---------------------------------------------|
| DARE  | Digital anorectal examination               |
| DARF  | Drug accountability record form             |
| DE    | Differentially expressed                    |
| DHHS  | Department of Health and Human Services     |
| DMC   | Data management center                      |
| DMR   | Differentially methylated region            |
| DNA   | Deoxyribonucleic acid                       |
| DSMB  | Data and Safety Monitoring Board            |
| ECOG  | Eastern Cooperative Oncology Group          |
| ELISA | enzyme-linked immunosorbent assay           |
| EMT   | Epithelial-mesenchymal transition           |
| ePRO  | Electronic patient-reported outcomes        |
| FCBP  | Female of childbearing potential            |
| FDA   | Food and Drug Administration                |
| FDR   | False discovery rate                        |
| FFPE  | Formalin-fixed paraffin-embedded            |
| FU    | Follow-up                                   |
| GO    | Gene Ontology                               |
| HIER  | Heat-induced epitope retrieval              |
| HIV   | Human immunodeficiency virus                |
| HPV   | Human papillomavirus                        |
| HRA   | High resolution anoscopy                    |
| HRQoL | Health-related quality of life              |
| HSIL  | High grade squamous intraepithelial lesions |
| IDB   | Investigational Drug Branch                 |
| IDE   | Investigational device exemption            |
| IHC   | Immunohistochemistry                        |
| IND   | Investigational new drug                    |
| IRB   | Institutional review board                  |
| IRC   | Infrared coagulation                        |  
| ITT    | Intent-to-treat                                |
|--------|------------------------------------------------|
| LAST   | Lower anogenital squamous terminology          |
| LCM    | Laser capture microdissection                  |
| LSIL   | Low grade squamous intraepithelial lesions     |
| KPS    | Karnofsky Performance Score                    |
| MDS    | Myelodysplastic syndrome                       |
| MOP    | Manual of Procedures                           |
| MSM    | Men who have sex with men                      |
| NCBI   | National Centre for Biotechnology Information  |
| NCI    | National Cancer Institute                      |
| NCTN   | NCI Clinical Trials Network                    |
| ODMC   | Operations and Data Management Center          |
| OHAM   | Office of HIV AIDS Malignancy                  |
| PCR    | Polymerase chain reaction                      |
| PGIC   | Participant global impression of change        |
| PIO    | Protocol Information Office                    |
| PR     | Partial response                               |
| PS     | Performance status                             |
| RACE   | Rapid amplification of cDNA ends               |
| RCT    | Randomized controlled trial                    |
| RLU    | Relative light units                           |
| RMA    | Robust Multi-Array                             |
| RNA    | Ribonucleic acid                               |
| SAE    | Serious adverse event                          |
| SCC    | Squamous cell carcinoma                        |
| SCJ    | Squamocolumnar junction                        |
| SIL    | Squamous intraepithelial lesion                |
| SIR    | Standardized incidence ratio                   |
| SISCCA | Superficially invasive squamous cell carcinoma |
| SOC    | System organ class                             |
| SOP    | Standard Operating Procedure                   |  
| STD   | Sexually transmitted disease            |
|-------|-----------------------------------------|
| TNF   | Tumor necrosis factor                   |
| TUA   | Treatment under anesthesia              |
| TZ    | Transformation zone                     |
| UADE  | Unanticipated Adverse Device Effect     |
| UCSC  | University of California, Santa Cruz    |
| UCSF  | University of California, San Francisco |
protocol synopsis
1.0 objectives 1.1 hypothesis

Treatment of anal high-grade squamous intraepithelial lesions (HSIL) will lead to a reduction of 75% of incident anal cancer compared with a population with anal HSIL that is observed without treatment.
protocol synopsis
1.2 primary objective

To determine whether treating anal HSIL is effective in reducing the incidence of anal cancer in HIV-infected men and women.
protocol synopsis
1.3 secondary objectives

1.3.1 To determine the safety of infrared coagulation, electrocautery, imiquimod, laser, and 5-fluorouracil treatments for anal HSIL.  
1.3.2 To assess the responsiveness (sensitivity to change) and clinical significance of the A-HRSI subscales by comparing change scores within groups of participants as defined by participant responses to the PGIC item.
protocol synopsis
1.4 exploratory objectives

1.4.1 Collect clinical specimens and data to create a bank of well-annotated specimens that will enable: a) Identification of viral factors in HSIL progression to cancer  
1.4.1.a.1 Determine the HPV type in cancer and compare to that of overlying HSIL and HSIL biopsies collected concurrently that did not progress to cancer.  
1.4.1.a.2 Determine the strain variant of HPV 16 in participants who progressed to anal cancer and compare to participants with HSIL biopsies who did not progress to cancer.  
1.4.1.a.3 Determine the HPV integration site in overlying anal cancer to that of HSIL overlying the cancer and HSIL biopsies collected concurrently that did not progress to cancer.  
b) Identification of host factors in HSIL progression to cancer 1.4.1.b.1 Perform gene expression array analysis comparing expression in anal cancer with HSIL overlying the cancer. Perform gene expression array analysis comparing expression in HSIL biopsies that progressed to cancer with non-progressing HSIL biopsies at other locations. Perform similar analyses comparing expression in HSIL biopsies that progressed to cancer with the same lesion at earlier time points prior to progression.  
1.4.1.b.2 Characterize genetic changes in anal cancers compared with HSIL
overlying the cancer. Characterize genetic changes in HSIL biopsies that progressed to cancer compared with non-progressing HSIL biopsies at other locations. Characterize genetic changes in HSIL
biopsies that progressed to cancer with the same lesion at earlier time points prior to progression.  
c) Identify host and viral biomarkers of progression from HSIL to cancer 1.4.2 Evaluate medical history and behavioral risk factors for HSIL progression to cancer.
protocol synopsis
1.5 quality of life objectives

1.5.1 Primary QOL Objective: To compare arms in terms of changes in physical symptoms and impacts from T2 to T3, adjusting for T1. Hypothesis: Participants assigned to the treatment arm will experience significant reductions in physical symptoms and impacts from T2 to T3 as compared to those in the active monitoring arm.  
1.5.2 Secondary QOL Objective: To compare arms in terms of changes in psychological symptoms from T3 to T4, adjusting for T1. Hypothesis: Due to potential uncertainty related to lack of treatment for those in the active monitoring arm, participants assigned to the active monitoring arm will experience significant increases in psychological symptoms from T3 to T4 as compared to those in the treatment arm.  
1.5.3 Exploratory QOL Objective: To assess of long-term HR-QoL changes in physical symptoms/impacts and psychological symptoms from T4 through the subsequent T5-T8 follow-ups overall and by arm.
protocol synopsis
1.6 ancillary study objectives

1.6.1 Determine the prevalence of SARS-CoV-2 detection in anal and oropharyngeal swabs among people living with HIV (PLWH) being screened for the ANCHOR study. Hypothesis: 10% of PLWH will have prevalent anal swab SARS-CoV-2 positivity, and will have concurrent oropharyngeal positivity.  
1.6.2 Determine the relationship in the ANCHOR screening population between prevalent anal SARSCoV-2 positivity, anal HPV infection, and anal high-grade squamous intraepithelial lesions (HSIL). Hypothesis: Anal SARS-CoV-2 positivity will be associated with higher quantities of anal HPV DNA and higher prevalence of anal HSIL among those who are highrisk HPV-positive.  
1.6.3 Determine the 6-month incidence of SARS-CoV-2 detection in anal and oropharyngeal swabs among participants with anal HSIL newly enrolled into the ANCHOR study.  
Hypothesis: Among PLWH enrolled into the ANCHOR study who were SARSCoV-2-negative at screening, 10% will have incident anal and oropharyngeal positivity at 6 months.  
1.6.4 Determine the relationship between prevalent or incident SARS-CoV-2 detection and regression of anal HPV infection or HSIL among participants newly enrolled into the 20 ANCHOR study and who are randomized to the monitoring arm.Hypothesis: In the monitoring arm of the ANCHOR Study, prevalent or incident anal SARS-CoV-2 detection will be associated with a lower rate of HSIL regression and clearance of anal HPV DNA than those who remain SARS-CoV-2-negative for 6 months.
protocol synopsis
2.0 background 2.1 anal cancer

2.1.1 Incidence of anal cancer in the general population and in high-risk groups Anal cancer is a growing problem in the United States. In the U.S. general population, the incidence of anal cancer from 2006-2010 was 1.5/100,000 among men and 1.9/100,000 among women (1). Updated estimates for the period 20112015 report the incidence at 1.15/100,000 among men and 1.93/100,000 among women (131). Compared with the general population, the standardized incidence ratio (SIR) of anal cancer is increased more than 100-fold among some risk groups of HIV-infected persons who are successfully treated with combination antiretroviral therapy (cART) (2). Among men in the general population, the incidence is highest among Black men, at 1.9/100,000 (3). The American Cancer Society projected that there would be 7090 cases and 880 deaths from anal cancer in 2013 (4). At the time the protocol was initiated, the most recent data with actual number of cases were available from 2009. There were 2,210 cases of cancer of the anus, anal canal, and anorectum among males and 3,624 cases among females. There were 302 deaths among males and 516 among females from these cancers. As of 2019, the rates of anal cancer were reported to be increasing at a rate of 2.7% per year from the period 2001-2015, with the number of cases from the period 20112015 at 9,598 among males and 18,683 among females; for the period from 20012016, there were 4,720 deaths among males and 7,391 among females (131).  
The incidence of anal cancer is largely concentrated among several groups wellknown to be at increased risk: men who have sex with men (MSM) (5), HIVinfected men and women (6), men and women immunosuppressed for reasons other than HIV infection, including solid organ transplant (7)(8), women with a history of HPV-related cancer or high-grade squamous intraepithelial lesions (HSIL) elsewhere in the anogenital tract (9), and those with a history of genital warts (10).  
Of all the groups listed above the highest incidence of anal cancer is among HIVinfected men and women, particularly MSM (2)(6).  
Overall it is estimated that the proportion of individuals with anal cancer who are also infected with HIV increased from 1980-1984 to 2001-2005, rising from 1.1% to 28.4% among males, and from 0% to 1.2% among females (11). From 19802005, the HIV epidemic had little impact on the trends in anal cancer among females, among whom the incidence has been increasing annually by 3.3%. However, the HIV epidemic has had a strong impact on the trends in anal cancer incidence among males, with an annual increase of 1.7% among men without HIV infection and 3.4% among men with HIV infection. The risk of anal cancer was elevated 52-fold in HIV-infected men who have sex with men (MSM), 32-fold in HIV-infected men, and 24-fold in HIV-infected women compared with the general population (11). Recent studies indicate an incidence of 131/100,000 among HIV-infected MSM in the U.S. from NA-ACCORD (6). The incidence of anal cancer was 46/100,000 among HIV-infected men other than MSM, and 30/100,000 among HIV-infected women. The incidence of anal cancer among HIV-infected individuals had also been reported outside the U.S. In the period from 2005-2008, compared with the general population, HIV-infected MSM from the French Hospital Database on HIV had a SIR for anal cancer of 109.8 (2). Other HIV-infected men had an SIR of 49.2 and HIV-infected women had an SIR of 13.1 (2). Among the four most common non-AIDS-defining malignancies, the largest increase has been in anal cancer (12). Several studies show an increase in the incidence of anal cancer since cART became available (11)(13)(14)(15). The relationship between risk of anal cancer, CD4 level, and complete versus incomplete HIV suppression is unclear. In the French study cited above, among patients who maintained a CD4 count above 500/mm3 for at least 2 years, a CD4 nadir below 200/mm3 was associated with a higher SIR for anal cancer than a CD4 nadir above 200/mm3 (2). Another report showed that being on cART with an undetectable HIV viral load 60% or more of the time was associated with reduced incidence of anal cancer compared with those who had an undetectable HIV viral load for less than 60% of the time (13). However, in that study the incidence of anal cancer was high, even among those receiving cART, and similar to the data reported in NA-ACCORD. Overall a cART-associated reduction in the incidence of anal cancer has not materialized (2). While it is possible that better control of HIV infection with earlier initiation of cART may moderate the increased risk of anal cancer, the long-term effect of cART on anal cancer risk has not yet been studied. Moreover, among HIV-infected individuals, the great majority of people currently living with HIV were begun on cART later in the course of their HIV infection than is currently recommended. A recent meta-analysis estimated that 1 in 377 HIV-infected MSM with anal HSIL progress every year to anal cancer since the introduction of cART (16), and since current cART has extended the life spans of HIV-infected individuals, this may result in a 10% lifetime risk of anal cancer or higher among HIV-infected MSM if nothing is done (17). Taken together, these data highlight two important points: HIV-infected men and women are disproportionately affected by anal cancer, and the impact of HIV infection on anal cancer will likely continue to increase. Second, most of the cases of anal cancer in terms of absolute numbers occur among HIV-uninfected men and women, and thus the results of the ANCHOR study will impact on both HIVinfected and HIV-uninfected populations. In the absence of active intervention, the number of cases of anal cancer will likely continue to grow in the general population in the U.S. with varying degrees of contribution by the at-risk groups described above. Data on the global incidence of anal cancer are not as reliable but it is certain that there are many thousands more cases and deaths from anal cancer each year world-wide. Combined with the possibility that anal cancer is preventable, the incidence of anal cancer is unacceptably high and calls for urgent intervention.
protocol synopsis
2.1.2 anal cancer may be

There are two approaches that may be used to prevent anal cancer. Primary prevention, in the form of vaccination against HPV infection may be useful to reduce infection with HPV, the underlying causative agent of anal cancer. Although there is great potential for HPV vaccination to reduce anal cancer in the long term, there are several limitations to this approach as described below. Secondary prevention, which consists of screening for, and treating HSIL prior to progression to cancer, is the approach needed for the many individuals who have already been exposed to HPV and who have developed HPV-associated precancerous lesions. Current cervical cancer prevention programs rely on both approaches, and in the long-term, it is likely that both will be needed for anal cancer prevention.  
2.1.3 A study of anal HSIL treatment in prevention of anal cancer is needed despite the availability of an HPV vaccine Like cervical cancer, most cases of anal cancer are caused by HPV 16 and HPV 18 (18). Vaccination with the quadrivalent and bivalent vaccine has been shown to reduce anal infection with HPV 16 and HPV 18 in both males and females naïve to those types (19)(20). Vaccination has also been shown to reduce the risk of anal HSIL in males (20). In the long-term, HPV vaccination may represent an excellent tool for reduction of anal cancer. Vaccination is currently routinely recommended for males (quadrivalent) and females (bivalent or quadrivalent) with a target of 1112 years of age, and up to 26 years of age for all women, and for men who are MSM, or HIV-infected/immunocompromised (21). Routine vaccination is recommended for other men up to age 21 years. However, the great majority of HIV-infected individuals currently at risk for anal cancer are older than 26 years and do not qualify for HPV vaccination. Even if the vaccine is made available to them, a high proportion will already have been exposed to HPV 16 and 18. Even among those who might benefit from vaccination, the impact of the vaccine has been reduced by poor uptake. Uptake among young women is limited at this time (22) with only 32% of eligible women receiving all three doses of the vaccine in 2010, and uptake among young men is even more limited. While herd immunity due to vaccination of females may contribute to protection against HPV even among those who have not been vaccinated, it is likely to be very limited among MSM. There are several reasons for the poor rates of HPV vaccination in the U.S. including varying levels of access, fear of HPV vaccine side effects, limited understanding of the benefits of HPV vaccination, and fear of vaccination in general. Globally the cost of the vaccine and lack of the necessary infrastructure to administer the vaccine are also major reasons. Finally, given the long period of time required for progression from cervical or anal HSIL to invasive cancer, it is expected that it will be decades before any reduction in cancer incidence is realized., even among those who have been vaccinated, it will likely be several decades before a vaccine-related reduction in anal cancer is observed and the full potential of the vaccine to reduce cancer incidence will most likely not be realized. In summary, the combination of low vaccine uptake among vaccine-eligible men and women means that the full potential of the vaccine to reduce cancer incidence will likely not be realized unless rates of uptake improve. Combined with the fact that most HIV-infected men and women are too old for vaccination or were exposed to HPV 16 and 18 before vaccination became available, millions of men and women remain susceptible to HPV 16- and HPV 18-related HSIL and cancer. For these individuals, secondary prevention in the form of identifying and treating HSIL may be the only option to reduce the risk of anal cancer. Determination of the efficacy of HSIL treatment to prevent anal cancer is therefore a current and public health concern for the foreseeable future for this target population.  
2.1.4 Similarities between anal and cervical cancer inform secondary prevention approaches to anal cancer prevention Cervical and anal cancer share many biological features. As described above, like cervical cancer, most cases of anal cancer are associated with HPV 16 or 18 infection (18). For both the cervix and the anus, the transformation zone is the primary target area for HPV due to the conjunction of two types of epithelium. In the cervix, these are the squamous epithelium of the exocervix and columnar epithelium of the endocervical canal. In the anus, the anorectal junction contains a transformation zone where the squamous epithelium of the anus is juxtaposed to the columnar epithelium of the rectum. HPV-associated changes in the cervix may be manifest as a spectrum of changes ranging from cervical low-grade squamous intraepithelial lesions (LSIL) to cervical HSIL. It is now well known that the precursor lesion to cervical cancer is cervical HSIL and the identification and removal of cervical HSIL before progression to cancer occurs is the basis for the success of the cervical cytology screening program. Similarly, anal squamous intraepithelial lesions (ASIL) span the same spectrum of changes as cervical lesions, and there is evidence that anal HSIL is the precursor to invasive anal cancer (23)(24) (25). In recognition of the similarity of HPV-associated anal and cervical disease, the American Society for Colposcopy and Cervical Pathology (ASCCP) and College of American Pathologists recently recommended standardization of terminology across all anogenital sites where HPV-associated cancers occur (LAST) (26). Prior to the introduction of cervical cytology screening programs, the incidence of cervical cancer in the U.S. was 40-50/100,000. Largely due to the success of cervical cytology screening, in which the cervical cancer precursor lesion, cervical HSIL, is sought and treated before progression to cancer occurs, the incidence of cervical cancer has declined to approximately 8/100,000 (27). Typically, an abnormal cervical cytology leads to referral for colposcopy, in which the lesions are visualized directly under magnification and with the aid of 5% acetic acid. The lesions are biopsied to establish the grade of the lesion, and HSIL is ablated using one or more different methods, most often loop electroexcision procedure in the U.S. Due to its success in reducing the incidence of cervical cancer, the cervical cytology screening program is considered to be one of the most successful cancer prevention programs ever implemented. Notably, however, the program was introduced prior to the era of evidence-based medicine, and its effect on reduction of cervical cancer only became apparent after following millions of women for several decades. While there are similarities between cervical and anal cancer, evidence must be developed as to whether anal cancer prevention is effective, to inform public health policy.  
2.1.5 A study of anal HSIL treatment in prevention of anal cancer is needed even though it is known that treatment of cervical HSIL is effective in prevention of cervical cancer Although anal cancer and cervical cancer are very similar, it cannot be assumed that treatment of anal HSIL will be effective in preventing anal cancer like treatment of cervical HSIL is effective in preventing cervical cancer. Compared with the cervix, a high proportion of anal lesions are large and multifocal, especially in HIV-infected individuals. The lesion recurrence rate and incidence of new lesions may be higher than is seen in the cervix in these groups. Compared with removal of cervical lesions using loop electroexcision procedure, complete removal of anal lesions can be comparatively difficult and may lead to more post-treatment discomfort given the need for regular bowel movements. Complete eradication of HSIL may therefore be more difficult in the anus than in the cervix. In addition, data on the efficacy of individual treatments for anal HSIL are far more limited than for treatment of cervical HSIL. While cervical cancer screening was adopted as standard of care prior to the era of evidence-based medicine, adoption of new medical practices such as screening and treatment of anal HSIL is increasingly requiring rigorous demonstration of the value of these practices. Although a growing number of groups around the U.S. have begun anal screening programs, but at this time only a small fraction of individuals at risk for anal cancer have had access to any form of anal screening and treatment for anal HSIL. With the exception of the State of New York, there are no formal public health guidelines recommending anal screening and treatment of anal HSIL. This is because no studies have yet been done to demonstrate that treatment of HSIL reduces the incidence of anal cancer. The American Society of Colorectal Surgery recently issued guidelines recommending that HSIL be treated, but based on low-quality evidence (28). The United States Public Health Service has indicated that evidence of the efficacy of HSIL treatment is needed before treatment of anal HSIL can be recommended (29). CDC STD guidelines for 2014 are being updated, and will state that "Some centers perform anal cytology to screen for anal cancer among highrisk populations (e.g. HIV-infected persons, MSM), followed by high-resolution anoscopy for abnormal cytologic results (e.g. ASC-US or worse). More evidence is needed concerning the natural history of anal intraepithelial neoplasia, the best screening methods and target populations, the safety and response to treatments, and other programmatic considerations before screening can be routinely recommended" (Park I, personal communication). Many insurance companies will not cover the cost of screening for anal HSIL or for treating it in the absence of guidelines establishing these services as standard of care. At the same time, standard of care guidelines require evidence that treatment of HSIL reduces the incidence of anal cancer. Conversely, if efficacy is not demonstrated, it is important that individuals with HSIL not be subjected to ineffective treatments that can confer medical harm and unnecessary cost to the healthcare system.
protocol synopsis
2.2 study agents and treatments

As noted previously, treatment of anal HSIL in HIV-infected men and women is more challenging than in HIV-uninfected individuals. This is because HSIL tends to be larger in size and number in HIV-infected individuals. They may also recur more often at the site of prior treatment, and incident lesions may develop more often at sites that were not previously shown to have HSIL (metachronous lesions). All of the treatments described below are used routinely by clinicians involved in treating HSIL. With the exception of treatment under anesthesia (surgical excision), which we expect to be necessary for only a very small percentage of patients with HSIL, each is readily available for use in the office setting by a wide variety of medical professionals, including MDs, nurse practitioners, and physician assistants. Thus, if the ANCHOR study shows that treatment of HSIL reduces the incidence of anal cancer, it will be possible to immediately implement the study findings.
protocol synopsis
2.2.2 infrared coagulation

Infrared coagulation (IRC) is an office-based procedure that uses the Redfield IRC 1900 or 2100, a therapeutic device to treat anal HSIL. It delivers short pulses of a narrow beam of visible and infrared light through a small contact tip applicator that is applied directly to the target tissue, transmitted down the rigid quartz glass of the light guide. The tungsten-halogen lamp (150 watts of power) is the light source. This light causes thermal coagulation that results in tissue necrosis. The depth of coagulation is determined by the total amount of energy delivered, which is adjustable via pulse duration setting (0.5 to 3.0 seconds in 0.1-second intervals) in repeated applications. The depth of tissue destruction is directly proportional to the duration of infrared impulses. The device is cleared by the Food and Drug Administration for use in the treatment of hemorrhoids, tattoo removal, chronic rhinitis, and anal condyloma. The procedure typically takes about 30 minutes to perform. It is generally well tolerated, and it is not uncommon for patients to go to work after the procedure.  
Overall the procedure is safe and can be performed by a wide variety of nonsurgeon medical professionals. It requires only local anesthetic injected directly into or around the lesions to be treated, and does not require smoke evacuation apparatus or protection. There may be occasional mild intra- and post-procedural pain, and bleeding for up to 2 weeks. There may also be mild textural changes of anal canal mucosa for several weeks post-procedure. The sequence of events is similar to that seen with cryosurgery: hemorrhagic blistering and necrosis of the treatment site followed by a shallow erosion and ulcer, and then healing over several weeks. There is a small risk of infection from the procedure. The risk of serious bleeding prompting emergency room evaluation is <1%. The PI of this protocol and his colleagues have performed thousands of IRC procedures, and most patients easily tolerate multiple procedures. Most published data show that on an individual lesion basis, the "cure" rate is high, defined as absence of HSIL in the treated area (30)(31)(32)(33) as well as when combined with excision under anesthesia for the patients with the most extensive disease (34). In these studies a therapeutic response is defined as absence of HSIL, and the remaining tissue may include normal tissue or LSIL. We expect that even if LSIL is present after IRC (or any treatment), that the main goal of treatment will have been attained, since there is no evidence that LSIL is a cancer precursor. In the largest study reported to date, the probability of curing an individual lesion after the first IRC ablation was 67% in HIV-infected men (30) and increased with additional ablations. Most recurrence was due to the development of metachronous lesions occurring in 82% of HIV-infected men after their first infrared coagulator treatment. At their last visit, 82% of HIV-infected MSM were free of HSIL. A similar result was obtained in AMC, in which the safety of IRC was studied in a multisite prospective cohort study performed by the AMC HPV Working Group (32).
protocol synopsis
2.2.3 electrocautery and surgical lasers

Electrocautery and laser ablation are ablative techniques with a long history of use in the operating room and the office for the treatment of HPV-associated anal lesions. A recent publication showed no statistically significant differences between electrocautery and IRC in their efficacy and safety profile (33). In the ANCHOR study, clinicians may use IRC, hyfrecator/electrocautery, and surgical lasers interchangeably for in office removal of anal HSIL. Legally marketed electrosurgical units and carbon dioxide lasers that are indicated for excision, coagulation, and/or ablation of tissue, and that are appropriate for use in dermatologic, general, and/or gastrointestinal surgical procedures may be used for in-office anal HSIL treatment on this protocol. The choice of therapy typically depends on the prior training of the clinician and the technique with which they have the most experience. Non-surgeons in the U.S. who treat anal HSIL were primarily trained by Dr. Palefsky and his group to use IRC, but those with a surgical background may prefer electrocautery. The main advantages of electrocautery are that it is faster than IRC to perform and the electrocautery probe tip can be more precisely applied to the lesion than the IRC probe tip, which is larger in diameter. The main disadvantage of electrocautery is that it generates a smoke plume, requiring protection from smoke for providers and patients in the room.
protocol synopsis
2.2.4 treatment under anesthesia

On this protocol, the term treatment under anesthesia (TUA) will be used to designate excision of HSIL lesions, performed by a surgeon on participants who are given spinal or general anesthesia. Procedures may be performed in an operating room as a surgical referral or in an office setting on participants with conscious sedation. The procedure will generally be performed using electrocautery devices or surgical lasers indicated for the excision, ablation, or coagulation of tissues, as described above. For TUA, selection of the medical devices to be used will be left to the discretion of the treating clinician. Combined with post-procedural IRC, it has been shown to be effective in treatment of HSIL for patients with disease too widespread to treat with targeted ablation therapy (34). TUA is usually required for only a small percentage of patients being treated for anal HSIL. These include 1) disease that is so extensive that it cannot be treated by any of the other methods described in this section, or 2) if a lesion is so large that it cannot be sufficiently biopsied in the office to establish or exclude a diagnosis of cancer to the satisfaction of the clinician. The disadvantage of TUA is that it is expensive compared with the other methods. In addition, by definition, TUA is only done when more extensive excision is required and patients typically experience more post-procedure side effects, including pain, bleeding, and infection after the procedure when compared with less aggressive treatment modalities. The costs of TUA will only be covered by the study with the QA committee's approval.
protocol synopsis
2.2.5 topical 5% fluorouracil cream

Topical treatment with 5-FU was first reported in 1962 for treatment of skin cancers following a report that systemic 5-FU induced regression of keratosis (35). It inhibits DNA synthesis by blocking the conversion of uracil deoxyribonucleotide to thymine deoxyribonucleotide, resulting in thymine deficiency and inhibition of cell division. Its most marked effect is on rapidly proliferating tissues where it can cause erythema and edema followed by erosion, ulceration, and necrosis. It has been used since the 1970s to treat HPV-related lower genital tract disease in women (36)(37). In one study of HIV-infected women, topical 5-FU was shown to be effective in preventing recurrence of cervical HSIL following loop electroexcision procedure (36). Most patients who use 5-FU exhibit local side effects in the form of pain or discomfort associated with epithelial ulceration. Moderate to severe side effects were reported by 48% of patients (38). In a subsequent randomized controlled trial, 27% of patients treated with 5-FU had grade 3 or 4 side effects (39). Topical 5-FU is typically used when disease is too extensive for IRC or electrocautery. It is given with the intent to clear as much of the lesion as possible, and while it is unusual for a large lesion to clear completely, it may be particularly useful to "debulk" a lesion to the point where it can them be treated with a targeted ablative approach such as IRC or electrocautery. Among 20 patients with extensive HSIL who were treated with topical 5-FU at UCSF, complete response was seen in only 3 and no response was seen in one, but 16 had a significant decrease in volume of disease to 25-50% that allowed for in-office IRC. At UCSF the protocol for use of topical 5-FU is for the patient to apply intra-anal or perianal 5-FU twice daily for 5 days, followed by a 9-day rest period (Jay N and Palefsky J, personal communication). This cycle can be repeated eight times. There are limited reports of use of topical 5-FU specifically for treatment of anal HSIL. 7 of 8 patients treated with topical 5-FU cream for perianal Bowen's disease applied twice weekly for 16 weeks had no evidence of Bowen's disease on biopsy at 12 months post-treatment (40). In another study using intra-anal 5-FU complete responses were seen in 12 of 34 patients with anal HSIL and partial responses with regression to LSIL in 8 of 34 patients (38). In a recently reported randomized controlled trial of 5-FU, imiquimod or electrocautery among 148 HIV-infected MSM with AIN (57% with HSIL) the response rate was 29, 39, and 48% respectively (39). Severe side effects were reported as 27, 43, and 18% respectively, and recurrence rates were 57, 25, and 17% at 6 months post-treatment. Of note the treatment regimens in this study were different from what we typically do and what is proposed for the ANCHOR study. The investigators used lower doses of 5-FU than we typically use, but their study participants also appeared to have more side effects than we have experienced with our patients.
protocol synopsis
2.2.6 topical 5% imiquimod cream

Like 5-FU, imiquimod is a topical agent that has typically been used when the extent of disease has been too large to allow for use of a targeted ablative modality such as IRC or electrocautery. Imiquimod has been used for treatment of external genital condyloma since 1997. It is a synthetic compound that exhibits antiviral activity by up-regulating the immune response, at least in part through toll-like receptors. It may lead to a Th1 cytokine response that activates HPV-specific cellmediated immunity and clearance of lesions. Imiquimod has been used for treatment of HPV-associated mucosal disease including vulvar (41) (42), penile (43)(44) and anal SIL (45). The advantage of imiquimod is that, like topical 5-FU cream it can be used to treat extensive disease that is too large for targeted ablation. It has a long safety record for treatment of condyloma, including in the setting of HIV infection. The disadvantages of imiquimod are side effects may be severe, particularly when patients are experiencing a robust clinical response. These may be local irritation or pain, or may be systemic flu-like symptoms. Another disadvantage is that there are relatively few randomized study data on the efficacy of imiquimod in treatment of perianal or anal disease. A randomized, controlled trial of 100 patients with anogenital warts assigned to 5% imiquimod cream or placebo for 16 weeks showed a low rate of complete response (11% drug vs. 6% placebo). 47% of patients in the imiquimod group reported greater than 50% reduction in total wart area vs. 20% in the placebo arm. Response rates were higher in HIV-uninfected patients compared with HIV-infected patients (62% and 31% respectively) (46). There are limited reports that demonstrate efficacy using imiquimod specifically for treatment of anal HSIL in cohort studies and case reports (47). In one study, 14 of 19 (74%) HIV-infected MSM had complete regressions after treatment with imiquimod (48). In a randomized, double-blind placebo controlled study of imiquimod for intra-anal HSIL, 4 of 28 participants on active drug resolved and 8 downgraded to LSIL, compared with only 1 of 25 in the placebo arm who had a spontaneous regression (49). Participants were treated with open-label imiquimod after completing the study, and overall 29 of 47 (61%) had sustained regression of anal HSIL. Our clinical impression over the years is that imiquimod has relatively limited value for treatment of anal HSIL, particularly in the setting of HIV infection where induction of immune response may be attenuated. However, we have not studied it formally in the U.S., and in light of the RCT data described above, the AMC HPV Working Group was recently approved by CTEP to perform a randomized trial of imiquimod vs. 5-FU vs. observation for extensive HSIL in HIV-infected men and women. We have also elected to leave imiquimod on the list of treatment modalities available to ANCHOR study clinicians but may change that pending the outcome of the AMC study described above or other study data as they become available.
protocol synopsis
2.3 study design and rationale

In this trial we will screen HIV-infected men and women for HSIL and if they meet the enrollment criteria for randomization, we will randomize them to treatment vs. active monitoring (AM) and follow them for up to five years after the last participant's date of randomization. Throughout the study we will be monitoring the incidence of invasive cancer in both arms and reporting regularly to a Data and Safety Monitoring Board (DSMB).  
We will also be collecting samples and clinical data that will allow us to perform the correlative science studies described in Section 9.0, which are focused on exploiting a unique opportunity to understand progression from HSIL to cancer.
protocol synopsis
definition of anal cancer

The anus is composed of the anal canal and the perianus. Proximally, the anal canal is bordered by the squamocolumnar junction (SCJ). This border is not static and anal squamous cell carcinoma (SCC) may occasionally be diagnosed proximal to the SCJ. These SCCs, diagnosed in the distal rectum, proximal to the SCJ, will be classified as anal cancers. Distally, the anus includes the perianus, defined as a circumferential area with a 5 cm radius from the anal verge, (after the buttocks have been retracted gently), and includes the squamous mucocutaneous junction (151). For a perianal SCC to be classified as an anal cancer, it must be part of a lesion (LSIL/HSIL/cancer) that is located wholly or partially within the perianus. If located outside the perianus, it must be contiguous with a LSIL or HSIL lesion that resides as least partially within the perianal region. An exception may be in the situation where an anal cancer is diagnosed outside the perianus, that was previously documented to be part of a lesion that was at least partially within the perianus, but at the time of cancer diagnosis a lesion within the boundaries of the perianus is no longer present due to prior treatment. Among women, the anterior border of the perianus is defined as the posterior half of the perineum. SCC located on the posterior half of the perineum are considered anal cancers even if they extend to the anterior, vulvar region.  
2.3.1 Rationale for selected approach and trial design To have an impact on standard of care guidelines, it is critical that a study of the value of anal HSIL treatment in prevention of anal cancer be designed to provide the highest quality of evidence. The highest quality evidence will be obtained in a randomized controlled trial (RCT). Such a trial would require HIV-infected men and women with biopsy-proven HSIL, randomized to a treatment or active monitoring arm, and followed for up to five years. Careful consideration has been given to alternative, less costly options for study designs that will yield the data needed to definitively define standard of care for anal cancer prevention. Least informative are small, non-randomized treatment studies of anal HSIL. There are few of these published to date. Two such studies showed that anal cancers occurred in patients whose anal HSIL was not treated, in contrast to patients whose HSIL was treated and who did not develop cancer (50) (52). Although encouraging, these small studies are subject to many potential biases and do not constitute definitive evidence of the value of anal HSIL treatment. A more robust study design would be an ecologic study, in which the rates of anal cancer at locations where HSIL is sought and treated are compared with locations where there is no such screening and treatment. There are two recent reports comparing the incidence of anal cancer in the San Francisco region, where the UCSF Anal Neoplasia Clinic has been treating patients since the early 1990s, with incidence reported from registries outside California. Individuals with anal cancer in the San Francisco region had a 39% lower mortality risk than those reported from registries outside California but there was no decrease in the incidence of invasive cancer in the San Francisco region (53). There are also data showing that while rates of anal cancer may not have decreased in the San Francisco area during this time, they may have increased more outside of California (54). Unfortunately, there are several limitations to these kinds of analyses with regard to inferences about the efficacy of treating anal HSIL in prevention of anal cancer. Methodologically-sound ecologic studies of anal HSIL treatment are not truly feasible for several reasons. First, screening and treatment for anal HSIL have begun on a small scale on an ad hoc basis in many centers with large populations of at-risk men and women. There is little standardization of clinical screening and treatment practices, and it is not possible to collect reliable data on treatment outcomes. Second, even in those locations where screening and treatment are performed, we estimate that only a small percentage of at-risk individuals have been screened to date and the effect of screening on reducing anal cancer incidence at these locations will be limited at best. This is true even in San Francisco, where we estimate that fewer than 10% of individuals with anal HSIL will have been treated, despite having one of the most active treatment programs in the country. Taken together, the data described above hint at the potential benefits of treating HSIL, but do not obviate the need for a RCT.
protocol synopsis
2.3.2. prevention of anal cancer

One option to reduce mortality from anal cancer would be to focus efforts on finding cancers as early as possible, instead of trying to prevent them by treating HSIL. In the absence of screening programs to identify anal cancer or anal HSIL in at-risk populations, most anal cancers are currently diagnosed when individuals present with anal pain, bleeding, or a mass. Occasionally asymptomatic cancers are detected on digital anorectal examination (DARE) or incidentally at procedures such as hemorrhoidectomy. Treating cancers when they occur, which is the current approach, has the advantage of not over-treating many individuals whose HSIL would never have progressed to cancer if left untreated. However, there are several reasons why focusing on diagnosis after cancer develops is a poor strategy. Many cancers found incidentally or because of symptoms will be diagnosed at more advanced stages. The survival rate declines substantially when the cancers are diagnosed at a more advanced stage (55). Even at stage 1, the most common stage at which anal cancer is diagnosed, nearly 30% of individuals die within 5 years. At more advanced stages, the mortality increases further, and when metastatic cancer is diagnosed (Stage IV) the 5-year survival rate is only 21%. Like cervical cancer, anal cancer may be preventable. An important reason to focus on cancer prevention rather than treatment of existing cancer is that while the standard anal cancer treatment, chemoradiation therapy (CRT), is often successful at the earlier stages of disease, it also leads to substantial morbidity (56). Acute toxicity is a major issue, particularly in HIV-infected patients, with one study reporting Grade 3 or 4 toxicity in nearly half of the patients (57). Chronic toxicity is also a major issue. Radiation therapy leads to radiation proctitis in a substantial proportion of patients, with anal pain and bleeding for years after successful therapy. Some require a colostomy for adverse late effects of CRT. The late effects likely result more from the radiation therapy rather than the chemotherapy. The symptoms related to treatment generally peak within the first 2 years, but radiation injury can develop after an interval of 5 to 10 years. Anal ulcers, fistulae, severe fibrosis, skin ulceration, and rectal stenosis may occur. There may be major issues with sexual function after CRT and there may be adverse psychosocial effects including depression. Radiation-induced pelvic and hip insufficiency fractures are also increasingly being recognized and reported, and are more common in women. Although HIV-infected individuals on cART are now commonly able to complete a full course of CRT, a study from the United Kingdom showed that CRT was associated with a reduction in CD4 level. The reduced CD4 level persisted after completion of therapy and may have contributed to the deaths of patients in remission (58). Currently we have no way to distinguish HSIL that will progress to cancer from HSIL that will never progress. This is a problem, since treatment of HSIL, while well tolerated, does have consequences in terms of acute morbidity and cost to the healthcare system. Further, HSIL is common among HIV-infected individuals, and a high proportion of HIV-infected men and women will be diagnosed with anal HSIL at some point if screening programs are implemented. Of note, however, even with the need to "over-treat" anal HSIL, screening for and treating anal HSIL is likely to be cost-effective compared with treating anal cancer. Several years ago Drs. Sue Goldie and Palefsky published formal cost-effectiveness studies of screening for and treating ASIL in HIV-infected and HIV-uninfected MSM (59)(60). Both of these studies showed these procedures to be highly cost-effective. Since that time, we believe that our treatment approaches have improved further, and that computation of cost-effectiveness today would yield even better numbers. Although the treatment of anal HSIL is not inexpensive, and may require multiple attempts and visits, anal screening and treatment is highly cost-effective because the risk of anal cancer is not evenly distributed throughout the population and we know who the high-risk groups are. A more recent study from the United Kingdom concluded that anal screening was not cost-effective for MSM, but the assumptions that went into their model were very different from those used in the Goldie papers, stemming from paucity of data and perhaps the very different cost structures of the U.K. and U.S. health care systems (61). Unlike cervical screening, which is recommended for all sexually active women, we only need to screen a subset of the population to have a substantial impact on the incidence of anal cancer. It is also possible that biomarkers of progression from HSIL to cancer can be identified, and this might reduce the number of HSIL lesions that need to be treated. This is one of the major aims of the correlative science component of the ANCHOR study and if successful, it will allow clinicians to target their therapy to only those HSIL at highest risk of progression. This would reduce unnecessary treatment and likely improve cost-effectiveness even further.
protocol synopsis
2.3.3 impact of preventing anal

In considering the value of preventing anal cancer, it is critical to consider both prevention of deaths and impact on quality of life among those who survive treatment of anal cancer. Simple treatments such as those that will be evaluated in the ANCHOR study to prevent development of cancer are far less morbid than treating invasive cancer (56). Clearly prevention of anal cancer through identification and treatment of HSIL would be preferable to waiting until cancers develop. If we can prevent most anal cancers, we will reduce the need for CRT in thousands of patients per year in the U.S., and prevent about 800 deaths per year. Worldwide we will reduce the need for CRT in many more thousands of patients, and likely prevent several thousand deaths per year. To address the issue of unnecessary treatment of anal HSILs that would not have progressed if left untreated, ANCHOR study has a robust correlative science component designed to identify biomarkers that have the potential to distinguish anal HSIL at high risk of imminent progression to cancer from anal HSIL that can be safely observed. Given the biological similarity to other HPV-related cancers, these data may be of value in prevention of cervical and HPV-related oral cancer, and may contribute to prevention of many hundreds of thousands, if not millions, of cases of cancer and deaths. The ANCHOR study correlative science program also has the potential to lead to new therapies for anal HSIL and cancer that would simplify treatment, and these too may potentially of value in prevention and treatment of cervical and oral cancer.  
2.3. Implementation of the ANCHOR study findings if the data support treating anal HSIL to reduce the risk of anal cancer
If the ANCHOR study shows that treatment of anal HSIL significantly reduces the incidence of anal cancer, it is likely that these results will quickly lead to standard of care guidelines. The ANCHOR study is a "strategy" trial, in which the strategy of treating HSIL is being tested for its ability to reduce the incidence of anal cancer. It is not a treatment trial of any one specific treatment modality. Thus, if the ANCHOR study shows that treatment of anal HSIL is effective in reducing anal cancer, standard of care guidelines will be adopted that use the most current information on how to treat anal HSIL, including the methods used in this study. If better methods become available in the future, the demonstration that treatment of anal HSIL reduces the risk of cancer will likely allow those newer treatments to be adopted as standard of care by showing their efficacy in treating HSIL. This is much easier and less expensive than needing to show efficacy in reducing the incidence Conversely, if the data from the ANCHOR study show that treating HSIL is not effective in reducing the incidence of cancer, we expect that dissemination of these findings will lead to a reduction and probably cessation of anal HSIL screening and treatment activities. If so, data from the ANCHOR study may still lead to an overall improvement in patient outcomes. Here it is possible that new progression biomarkers and therapeutic approaches identified in the ANCHOR study correlative science program may allow us to identify and treat anal cancers much earlier than is the case currently. This in turn may lead to successful treatment of these cancers with much less morbidity than CRT. It is very possible that the data will also be of value for prevention and treatment of cervical and oral cancer. If treating HSIL is effective in reducing the incidence of anal cancer, then methods to identify those with anal HSIL through screening of at-risk individuals will become standard of care as well. As with treatment, there are several approaches that are currently available, but newer screening methods are being tested and others will be tested in the future. Correlative science studies enabled by the ANCHOR study as well as several others currently in progress will likely lead to optimization of screening strategies for anal HSIL, to the identification of those patients with HSIL who are at increased risk of progression to anal cancer, and potentially to new treatment approaches to HSIL and cancer.  
2.3.5 The impact of the ANCHOR study for setting public health policy extends well beyond HIV-infected men and women
In addition to patients with HIV infection, those with a history of genital warts, or signs of perianal HPV-related lesions, whether warts or HSIL, may also potentially benefit from screening. With the growing incidence of anal cancer among the general population of women, we may reach a point where screening larger numbers of women who do not fit into any of these groups (e.g., women reporting a history of anal intercourse), may also become useful, but this will require further study. Although the non-HIV-infected groups listed above are not eligible for the ANCHOR study for several reasons outlined in Section 2.3.7, we also expect that the standard of care guidelines adopted for HIV-infected men and women will apply to the non-HIV-infected groups as well. Anal HSIL in HIV-infected men and women is harder to treat than in HIV-uninfected men and women for a number of reasons, including multifocal lesions, large lesion area, and high risk of HSIL recurrence after treatment. We believe that if the ANCHOR study shows that treatment reduces the risk of anal cancer in HIV-infected individuals, then the results will be generalizable to other groups in whom treatment of HSIL will be less challenging. Since most cases of anal cancer in the general population are occurring in HIV-uninfected individuals, it is clear that the data from ANCHOR study will have an impact on populations at risk for anal cancer that extend well beyond those with HIV infection.
protocol synopsis
2.3.6 the impact of the

The ANCHOR study is also designed to have wide impact through its correlative science component. Multiple samples will be collected from patients during the trial, and we plan to analyze them through a series of separately funded correlative studies. The ANCHOR study is unique because it focuses on prevention of progression from HSIL to cancer. A study such as this is not currently possible at any other anatomic site at which HPV-associated cancer develops. A study of progression to cervical cancer is not possible because it is currently standard of care to treat cervical HSIL. A study of progression to oral cancer is not possible because it is difficult to reliably identify oral cancer precursor lesions for treatment. The ANCHOR study not only offers the opportunity to determine the efficacy of HSIL treatment to prevent cancer, but careful follow-up and sample collection from those in the active monitoring arm will allow us to better understand the molecular pathogenesis of progression to cancer. This may lead to new approaches to treating HSIL to prevent progression to cancer and potentially cancer itself. The study also provides the opportunity to identify biomarkers of progression to cancer. Given the strong biological similarities between anal, cervical and oral cancer it is very possible that biomarkers and treatments identified in the ANCHOR study will be applicable to cervical and HPV-associated oral cancer. Biomarkers of progression from cervical HSIL to cervical cancer would be of great value in resource-limited settings where the cost of treatment could be reduced if there were ways to identify those at highest need of intervention. This is often the case in developing countries that lack organized cervical cancer screening programs. In developed countries, this information could be valuable to identify those at highest need of close follow-up after treatment of cervical HSIL. Recurrence after treatment of cervical HSIL is particularly common in HIV-infected women (62) and this is just one group that might benefit from use of these biomarkers. Development of better treatments to reduce the incidence of HPV-related cancers regardless of the anatomic site at which they occur, or better biomarkers to identify those at risk, and who most need treatment of HSIL, has the potential to save hundreds of thousands, if not millions of lives. In summary, the ANCHOR study will have a major impact in several ways: 1) If treatment of HSIL is shown to be effective in reducing the incidence of anal cancer, it will establish standard of care guidelines for treatment of HSIL in at-risk populations. The results will be applicable to both HIV-infected and HIVuninfected at-risk populations; 2) Information obtained on molecular pathogenesis of progression from anal HSIL to anal cancer, and development of biomarkers of progression may lead to an ability to target those with anal HSIL most in need of treatment, and better methods treating anal HSIL to prevent anal cancer, and 3) the results of ANCHOR will likely extend to cervical and HPV-associated oral cancer.
protocol synopsis
2.3.7 rationale for eligibility criteria

The ANCHOR study includes only HIV-infected men and women for several reasons. First, this is the group with the highest incidence of anal cancer. Second, since the incidence of cancer is highest in this group, we expect to obtain the necessary number of cases of anal cancer in the active monitoring arm with the smallest number of participants possible, and over the shortest amount of time possible. Third, given the challenges associated with treating HSIL in this group (multifocal lesions, risk of HSIL recurrence after treatment) we believe that if the study shows that treatment reduces the risk of anal cancer in HIV-infected individuals, then the results will be generalizable to other groups in whom treatment of HSIL will be less challenging. In addition, performing a RCT like ANCHOR study in HIV-uninfected individuals would not be practical since such a study would require many more participants who would need to be followed for a substantially longer period of time. It would also be difficult to generalize the results of any study in HIV-uninfected individuals to HIV-infected individuals. We also have elected to study only HIV-infected men and women 35 years or older. This is meant to enrich the study population at risk for cancer since anal cancer occurs only rarely under this age even among HIV-infected individuals. Fewer than 1% of anal cancers occur under the age of 35 years (1). Finally, our goal is to recruit to this study at locations around the U.S. that will allow us to enroll participants whose racial/ethnic background mirror the current demographics of the HIV epidemic, to ensure the generalizability of the results to the HIV-infected population into the future. We have considered other potential methods to enrich the study population for risk of cancer such as HPV 16 or 18 positivity as an enrollment criterion but other HPV types may also play a role in anal cancer, particularly in the HIV setting, and we want to ensure that our results are as generalizable as possible. Although HPV 16 is the predominant HPV type in anal cancers overall (18), little is known about the distribution of HPV in anal cancers among HIV-infected men and women. Recent data suggest that HPV 16 may play a less prominent role in cervical cancers of HIV-infected women compared with HIV-uninfected women (63)(64), and it will be important to know whether this is also true of anal cancers. In one study from Germany, all five squamous cell carcinomas (SCCs) of the anal canal contained HPV 16 compared with only one of four anal margin SCCs. HPV 31, HPV 33 and HPV 68 were found in the other three anal margin SCCs) (52). We do not plan to use CD4 level, HIV viral load, history of cART, and nadir CD4 level as entrance criteria, since our own data show that individuals on cART have a wide range of CD4 levels at the time of anal cancer diagnosis (25). A high proportion of HIV-infected individuals are on ART and thus use or lack of use of cART also would not be a useful entrance criterion. Other than the recent publication using VA data suggesting that having more time with an undetectable HIV viral load was protective against developing anal cancer (13), there are insufficient data to use HIV viral load as an entrance criterion. It would be difficult to reliably quantify a prospective participant's past viral load control, and regardless of what it was at the time of study entry, we would feel ethically obliged to work with the participant and their HIV care provider to maximize the likelihood that they have the best viral load control possible.
protocol synopsis
2.3.8 equipoise in the treatment

We have had many discussions with specialists in medical ethics and with many members of the HIV community. We believe that there is equipoise in the treatment and active monitoring arms of the ANCHOR study. Participants in the active monitoring arm are at risk of progressing to anal cancer. However, we do not know that treatment of anal HSIL is effective to reduce anal cancer, and individuals in the treatment arm may be at similar risk, hence the need for the ANCHOR study. If HSIL treatment is shown to be ineffective in prevention of incident HSIL, then participants in the active monitoring arm are being spared treatments that may be causing unnecessary morbidity. We have designed the study to minimize the risk of harmful outcomes associated with developing invasive cancer for ethical reasons and to maximize the likelihood of study retention. For this reason, we are following participants in both arms very closely, i.e., at 6-month intervals or shorter. With close follow-up, it is possible (if not probable) that if a cancer does develop, it will be an early cancer. Detecting cancers early is likely to be advantageous since treatment is usually successful at the early stages, and is less successful at later stages. Cancers that occur while still defined as superficially invasive squamous cell carcinoma (SISCCA) (26) may be treatable with local excision. Local excision is not standard of care for intra-anal SISCCA but this is a research question that will be addressed in a study under investigation by the AMC HPV Working Group (participants in the ANCHOR study who are diagnosed with SISCCA may be referred to that study if it is still recruiting). Local removal could potentially spare participants the side effects associated with CRT. However, in the informed consent process, we will be explicit with the participants that despite the frequent monitoring in the study, those who develop cancer may need CRT with its associated morbidity, and they may also die of their anal cancer. It is very possible that in the absence of the ANCHOR study, many individuals who agree to participate and who are randomized to the active monitoring arm would not have undergone any form of screening- screening is not currently standard of care and is not available to most at-risk individuals. If they were going to develop cancer, they would have done so anyway, outside the context of any study and these individuals would eventually present with symptoms from their cancer. The cancers may be diagnosed at a later stage than if they were being monitored in the ANCHOR study, would more likely require CRT and they may have increased risk of death. Being in the active monitoring arm therefore offers the possibility of earlier diagnosis of cancer than if they were not in the study at all, a better chance of survival and very possibly fewer side effects of cancer treatment. Being in the treatment arm offers the possibility of reduced risk of developing anal cancer compared with being in the active monitoring arm. However, as noted above, we do not yet know if this is the case, and it is possible that individuals in the treatment arm will be undergoing a series of procedures with little or no clinical benefit. Many of the treatments described above lead to a high clearance rate per lesion, but patients may still have a focus of anal HSIL somewhere even after successful treatment of a given lesion because of metachronous disease. We will therefore allow ongoing treatment of HSIL throughout the course of the study. Although the HSIL treatment procedures that we will use in the ANCHOR study are generally well tolerated, participants in the treatment arm may experience treatment-associated side effects, including post-procedure pain or discomfort, bleeding and although rare, infection or stenosis. The DSMB will review data including progression rates to cancer in both arms on a regular basis (e.g., 6 months, and not less than once per year) and will discontinue the study if the data indicate that it is no longer ethical to continue. The DSMB will also monitor study data relevant to the assumptions so that necessary changes can be made to accrual strategies, eligibility criteria, sample size, or other study parameters in a timely manner. Interim analyses are described in Section 10.5.2. In summary, participants in both arms who develop anal cancer may need CRT, and may be at risk of death due to anal cancer. It is not known if this risk is different between the two arms. Participants in both arms may also benefit from early diagnosis of anal cancer and may be spared the side effects of CRT. Some might benefit from being in the study compared with not being followed at all, which is currently the case for most at-risk individuals. Finally, if the study hypothesis is rejected, i.e., that treatment of HSIL is not effective to reduce the incidence of anal cancer, then the participants in the active monitoring arm will have been spared the morbidity associated with treatments of those participating in the treatment arm.
protocol synopsis
2.3.9 feasibility of recruitment to

In preparation for the ANCHOR study, we performed a series of studies examining willingness to enroll in the study. Funded by NIH, we performed focus groups in ten different U.S. cities comprised of HIV-infected men and women. In each city we obtained data from several groups, individually composed of HIV-infected White, Black and Hispanic men or women. Our goal was to assess the feasibility of performing the ANCHOR study among the demographic groups that currently comprise the HIV epidemic. Given that we expect to recruit to the ANCHOR study in 10 sites spread around the United States where the HIV epidemic is prominent, we sought to assess willingness to participate across a wide geographic and demographic spectrum of potential participants. We assessed willingness to participate in a trial with the same design as ANCHOR in an ethnically diverse sample of HIV-infected men (n=202) and women (n=39) in 10 US cities. Focus group participants were screened for eligibility (35 or more years of age and HIVinfected) by telephone and assigned to one of four focus group types based on ethnicity and gender (Black, Hispanic or Latino, and White men, and a multi-ethnic group of women). Two moderators traveled to each of the ten cities to conduct the focus groups following a semi-structured guide. The Hispanic groups were conducted primarily in Spanish, primarily in English or both, depending on participant preference. We also performed an internet-based survey of 257 HIV-infected men and women in 20 cities. The online survey targeted the same population living in the same 10 cities used for the focus group portion of the study, plus 9 additional US cities (Boston MA, Minneapolis MN, Omaha NE, Philadelphia PA, Sacramento CA, San Antonio TX, San Diego CA, Tampa FL, and Washington DC) and one Canadian city (Vancouver BC). The study was conducted using SurveyGizmo and participants were recruited through flyers mailed to AIDS service organizations and HIV clinics. We also directly surveyed HIV-infected men to determine their willingness to participate in study with the ANCHOR study design after being given a diagnosis of anal HSIL. In summary, most HIV-infected individuals surveyed expressed an interest in participating in the ANCHOR study even with a 50% chance of being randomized to either arm. In so doing they would be supported by their HIV primary care provider. Survey respondents mostly preferred being in the treatment arm, but with appropriate support, would likely stay in the active monitoring arm. Among the 20 participants who were surveyed after being given a diagnosis of HSIL, most expressed a willingness to be randomized even after having undergone procedures very similar to the ANCHOR study screening protocol and after having come to the clinic fully expecting to be treated for their HSIL. In the worst-case scenario, individuals may choose not to continue in the study after their screening visit, and if that is the case, we will screen additional individuals to reach our recruitment target of 5058 randomized men and women. Taken together, these data give us confidence that we will be successful in recruiting to the ANCHOR study as described in the schema, and that we will be able to successfully retain participants in both arms with sufficient ongoing education and support.