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2.2 screening evaluation

2.2.1 Screening activities performed prior to obtaining informed consent Minimal risk activities that may be performed before the subject has signed a consent include the following:  
Email, written, in person or telephone communications with prospective subjects Review of existing medical records to include H&P, laboratory studies, etc.  
Review of existing MRI, x-ray, or CT images Review of existing photographs or videos Review of existing pathology specimens/reports from a specimen obtained for diagnostic purposes.
2.2.2 Screening activities performed after a consent for screening has been signed The following activities will be performed only after the subject has signed the consent for this study for screening. Assessments performed at outside facilities or on another NIH protocol within the timeframes below may also be used to determine eligibility once a patient has signed the consent. Performed at any time prior to start of study therapy Archival tumor sample for NCI LP confirmation of diagnosis: The mesothelin expression analysis will be performed by the NCI Laboratory of Pathology at the time of histological confirmation of non-squamous non-small cell lung cancer. This must be done prior to subject enrollment to trial and treatment. A block of primary tissue (or 5-10 unstained sections on charged slides) from the time of diagnosis will be required from each patient. Tissue blocks from a known recurrence will be accepted if original tumor samples are unavailable. Referring institutions will send the tumor block or 5-10 unstained sections on charged slides to CCR/NCI for correlative studies and confirmation of diagnosis. A fresh biopsy or tumor effusion sample will be collected if archival tumor tissue is not available.  
Confirmation of EGFR, ALK1, ROS1 and KRAS mutation status in patients with NSCLC
In cases where archival tumor tissue is unavailable, a fresh biopsy sample will be taken and sent to the NCI Laboratory of Pathology to screen for EGFR, ALK1, ROS1 and KRAS genes using the following assays:  
| genes using the following assays: Assay Mutation Being   | Assay Description                                                                                                                                                                                                                                                                                 | FDA Cleared for   |                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                               |     |
|----------------------------------------------------------|---------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|-------------------|---------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|-----|
| Tested                                                   | This Use?                                                                                                                                                                                                                                                                                         |                   |                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                               |     |
| 1                                                        | FISH ALK                                                                                                                                                                                                                                                                                          | ALK               | LSI ALK (anaplastic  lymphoma kinase) Break  Apart (BA) DNA FISH test  (in-house): This is an FDA  approved companion  diagnostic test designed to  detect rearrangement of ALK  gene via FISH in FFPE tumor  samples from patients with  NSCLC and other tumor  types. The Dual Color, Break  Apart Rearrangement Probe  from Abbott is designed to  detect the known 2p23  rearrangements of ALK due to  pericentric inversion or  translocation events in lung  cancer. It will detect most of  the ALK gene rearrangements  regardless of fusion partner. | Yes |
| 2                                                        | TruSight  Oncology 500  Assay  (TSO500)                                                                                                                                                                                                                                                           | EGFR, ALK and     | TSO500 gene panel is                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                          |     |
| ROS1                                                     | designed to provide full gene  coverage for 523 genes. This  assay is intended to detect  pathogenic or potentially  pathogenic variants relevant to  cancer and does not  distinguish somatic variants  from germline variants. It will  be able to detect most of the  ROS1 rearrangements with | No                |                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                               |     |  
| known and unknown fusion  partners. The limit of detection (LOD)  for SNVs and indels is 5%  variant allele frequency as  determined by analytical  sensitivity testing using  reference samples containing  approximately 200 known  common variants. However, it  is important to note that the  LOD for this assay, at any  particular nucleotide, is  correlated with the sequencing  coverage obtained at that  locus. The limit of detection  of the fusions covered by this  assay is undetermined but  requires a minimum 10%  tumor cells.   |
|-------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|  
Performed within 28 days prior to start of study therapy or, if applicable, second course History and physical exam Vital signs including pulse oximetry  
ECG
Echocardiogram CT scan of chest, abdomen and/or pelvis and areas of known or suspected disease involvement; MRI* may also be performed when appropriate. Additional imaging may be done as clinically indicated. (If patient being assessed for Second Course, please see Appendix D)
18F FDG-PET/CT scan Urinalysis
*NOTE: MRIs done in this study will involve the use of the contrast agent gadolinium, unless contraindicated. The risks associated with MRIs and contrast are discussed in the consent form.
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performed within 10 days prior

CBC with differential, Acute Care Panel (sodium, potassium, chloride, bicarbonate, creatinine, glucose, BUN), Hepatic Panel (alkaline phosphatase, AST, ALT, total bilirubin, direct bilirubin), Mineral Panel (albumin, calcium, magnesium, phosphorus), creatine kinase, C-reactive protein, Coagulation (PT, PTT, fibrin degradation products), lactate dehydrogenase, Thyroid (TSH, free T4, total T3)
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performed within 72 hours prior

Urine or serum hCG in women of childbearing potential (must be repeated on C1D1 (pretreatment) if more than 72 hours have passed since screening assessment)
2.3 PARTICIPANT REGISTRATION AND STATUS UPDATE P**ROCEDURES**
Registration and status updates (e.g. when a participant is taken off protocol therapy and when a participant is taken off-study) will take place per CCR SOP ADCR-2, CCR Participant Registration & Status Updates found here.
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2.3.1 screen failures

Screen failures are defined as participants who consent to participate in the clinical trial but are not subsequently assigned to the study intervention or entered in the study. A minimal set of screen failure information is required to ensure transparent reporting of screen failure participants, to meet the Consolidated Standards of Reporting Trials (CONSORT) publishing requirements and to respond to queries from regulatory authorities. Minimal information includes demography, screen failure details and eligibility criteria. Individuals who do not meet the criteria for participation in this trial (screen failure) because of a laboratory abnormality (except for negative mesothelin) may be rescreened.  
2.4 TREATMENT ASSIGNMENT P**ROCEDURES**
2.4.1 Cohorts:  
| Number         | Name                              | Description                                              |
|----------------|-----------------------------------|----------------------------------------------------------|
| 1              | Mesothelin expressing             | Up to 21 evaluable NSCLC subjects previously treated     |
| NSCLC subjects | with standard therapies           |                                                          |
| 2.4.1.1        | Arms                              |                                                          |
| Number         | Name                              | Description                                              |
| 1              | LMB-100 +                         | LMB-100 administered in first two cycles + pembrolizumab |
| Pembrolizumab  | administered in subsequent cycles |                                                          |  
2.4.2 Arm Assignment Patients in cohort 1 will be directly assigned to arm 1.
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3 study implementation 3.1 study

This is an open-label, single center phase II study of LMB-100 followed by pembrolizumab in patients with advanced non-squamous non-small cell lung cancer who must have progressed on standard therapies including an immune checkpoint inhibitor.  
Patients will receive LMB-100, 140 mcg/kg, on days 1, 3 and 5 of a 21-day cycle for the first two cycles.  
Subjects will receive pembrolizumab monotherapy (200 mg) on day 1 of each subsequent 21-day cycle for up to 2 years or until disease progression on pembrolizumab, or intolerable toxicity, whichever occurs first.  
Patients who are progressing after cycle 1 of LMB-100, will be allowed to undergo early restaging scans to evaluate tumor response. If there is either radiological (RECIST) or clinical progression after cycle 1, patients will be treated antecedently with pembrolizumab starting cycle 2. In this case LMB-100 will be discontinued and will not be given at cycle 2.  
Pembrolizumab dosing will not be initiated until LMB-100 related toxicities have resolved to grade 1 or better; however, if the initiation of pembrolizumab is delayed for more than 4 weeks, the patient will be removed from study therapy.  
All participants who stop study treatment with stable disease or better may be eligible for an additional 17 cycles of pembrolizumab if they progress after stopping study treatment from the initial phase.  
Response evaluation will be done every 6 weeks Optional tumor biopsies for research will be performed at baseline, at the end of cycle 2 of LMB-100, after second cycle of pembrolizumab, to evaluate changes in the tumor immune microenvironment following treatment with LMB-100 and pembrolizumab. These biopsies are optional, and a fresh biopsy or tumor effusion sample will be collected if archival tumor tissue is not available. In the event that patients get one cycle of LMB-100, optional tumor biopsies for research will be performed at baseline, at the end of cycle 1 of LMB-100 and after second cycle of pembrolizumab.
* Note: A weight-based dose cap will be applied. LMB-100 dose for patients weighing more than 100 kg will be calculated as if they weigh 100 kg.  
3.1.1 Schema and dose schedule
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3.1.2 second course of treatment

All participants who stop study treatment with stable disease or better may be eligible for up to an additional 17 cycles (approximately 1 year) of pembrolizumab treatment if they progress after stopping study treatment from the initial treatment phase. This retreatment is termed the second course phase of this study and is only available if the study remains open and the participant meets the following conditions:
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either

Stopped initial treatment with study treatment after attaining an investigator-determined confirmed CR based on RECIST 1.1, and o Was treated with at least 8 cycles of study treatment before discontinuing treatment, and o Received at least 2 treatments with pembrolizumab beyond the date when the initial CR was declared
OR  
Had SD, PR, or CR and stopped study treatment after completion of 35 administrations
(approximately 2 years) of study treatment for reasons other than disease progression or intolerability
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and

Experienced an investigator-determined radiographic disease progression by RECIST 1.1 after stopping initial treatment, and o No new anticancer treatment was administered after the last dose of study treatment, and o The participant meets all of the safety parameters listed in the inclusion criteria and none of the safety parameters listed in the exclusion criteria, and o The study is ongoing
An objective response or disease progression that occurs during the second course phase for a participant will not be counted as an event for the safety and efficacy endpoints of the study.
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3.2 drug administration 3.2.1 lmb-100

The qualified health care professional responsible for dispensing the study drug will prepare the correct dose according to the cohort allocation of each patient. LMB-100 (see section 3.1 for dose) will be given as an IV solution on Days 1, 3, and 5 (every other day) of a 21-day cycle for two cycles. LMB-100 must be administered in a hospital or clinic equipped for IV chemotherapy. Full emergency resuscitation facilities should be immediately available, and patients should always be under close supervision of the investigator or delegate. The compatibility and stability of the active ingredient was tested under simulated preparation/administration conditions. 3.2.1.1 Instructions 1. Visually inspect the LMB-100 drug product prior to dose preparation. The solution should be colorless to brownish and may contain a small amount of translucent-to white amorphous LMB-100 particulates. Do not use if other particulate matter is seen or if the study drug is discolored. Do not shake the vial to mix.  
2. Use an appropriately sized syringe and needle to draw up the required volume of LMB100 (undiluted drug) plus an additional 3 mL (overfill for priming the syringe extension line) from the vial.  
3. Attach a Smiths Medical Ultra™ Small Bore Extension Set (REF \#MX448HL60) to the syringe and a Smith medical 0.2-micron filter set (MX448HF) to the end of the extension set, then prime close to the distal end. Purge all air from the syringe and line. Cap with a sterile Spiro cap before dispensing.  
3.2.1.2 Administration Instructions 1. Intravenous (IV). Do not administer as IV push or bolus.  
2. LMB-100 is administered either peripherally or centrally through a patient's vascular access device. If there is no pre-existing central vascular access device (VAD) and peripheral access is inadequate, a central access device will be installed.  
3. Undiluted LMB-100 (1 mg/mL) will be transferred to a disposable syringe and administered by IV infusion using a syringe pump. LMB-100 will be diluted in-line 1:10 with 0.9% Sodium Chloride Injection, USP immediately prior to administration.
4. To accomplish this, a side flow with 0.9% Sodium Chloride Injection, USP, must be applied. A 4-way stopcock (alternatively two 3-way stopcocks joined end-to-end, or an alternative manifold with at least two inlets and one outlet ports that can be opened simultaneously) should be positioned at the patient's VAD to allow simultaneous infusion of LMB-100 and 0.9% Sodium Chloride Injection, USP. The infusion rate of the 0.9% Sodium Chloride Injection will be 9-times the hourly rate of LMB-100.  
5. LMB-100 will be filtered inline during administration with a 0.2-0.22micron filter.  
  
For the first infusion, venous access (via peripheral IV or central line) should remain accessible for 2 hours after the end of infusion or longer, if necessary, during the remainder of the hospital stay. Duration of use of venous access will comply with CC Nursing policies and procedures. If no infusion related symptoms occur during this time, the infusion line may be removed or deaccessed. For subsequent infusions and if no IRR has been reported, venous access should remain in place for at least 30 minutes from the end of the infusion. If no adverse events occur during the 30 minutes, the infusion line may be removed. During the infusion, vital signs (including, if possible, supine diastolic and systolic blood pressure, pulse rate, and temperature) must be monitored pre-infusion and every 15 minutes (± 5 minutes) until the end of the infusion. Following the completion of the infusion, vital signs should be checked every 30 minutes (± 10 minutes) for the first 2 hours and once at 4 hours after infusion completion. Vital signs during the infusion are not required to be captured in the eCRF unless abnormalities are observed. LMB-100 drug product (DP) should be administered diluted using a side flow set-up at 1:10 (0.1 mg/mL DP). In order to not compromise drug product physico-chemical stability, the dilution with 0.9% NaCl should be done **in line**, immediately prior to administration of the DP. The infusion duration should be 30 minutes (- 5 minutes, +10 minutes); however, the duration can be increased at the discretion of the investigator based on the total dose and volume to be administered and the patient's physical condition. Syringe preparation and infusion duration should not exceed a maximum of 4 hours. In case of any adverse events related to the infusion, please refer to the specific recommendation described in section **3.3.1**. 3.2.2 Pre-medications for Patients Receiving LMB-100 Due to the prevalence of infusion related reactions (IRRs) seen in the previous study of LMB-100, all patients will be predicated 30-60 minutes (+ 30 minutes) prior to each LMB-100 administration with the following medications (**Table 5**):  
o Diphenhydramine 25-50 mg PO or IV o Famotidine 20 mg PO
o Acetaminophen 650 mg PO
(See section **3.3.1** for complete instructions on response to IRRs)
Note: An alternative histamine H2 antagonist may be substituted for famotidine, if the preferred pre-medication is unavailable.  
| Dose (mg)        | Route   |              |
|------------------|---------|--------------|
| acetaminophen    | 650     | Orally       |
| famotidinea      | 20      | Orally       |
| diphenhydramineb | 25-50   | Orally or IV |  
a An alternate histamine H2 antagonist or famotidine IV are allowed if the preferred pre-medication is unavailable b or alternative antihistamine at an adequate dose
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3.2.3 pembrolizumab

Pembrolizumab 200 mg will be administered as a 30-minute IV infusion every 3 weeks = 1 cycle. Pembrolizumab will be administered on day 1 (± 3 days) of each cycle starting with cycle 3 (if conditions in section **3.3.2** are met) for up to 2 years unless criteria for removal from therapy (**3.6.1**) are met. Patients who are progressing after cycle 1 of LMB-100, will be allowed to start pembrolizumab from cycle 2 onwards. Note: As indicated in section 3.1, in certain circumstances the total duration of pembrolizumab treatment may exceed the 2-year limit. Every effort will be made to target infusion timing to be as close to 30 minutes as possible. However, given the variability of infusion pumps, a window of -5 minutes and +10 minutes is permitted (i.e., infusion time is 30 minutes: -5 min/+10 min). Vital signs (blood pressure, pulse, respiratory rate, temperature) will be measured pre and post infusion. Pembrolizumab may be administered on an outpatient basis. The Pharmacy Manual contains specific instructions for the preparation of the pembrolizumab infusion fluid and administration of infusion solution. Only if medically warranted in response to an AE will the Investigator modify the infusion parameters (decrease the rate of infusion, interrupt the infusion, or reduce the dose volume infused).
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3.3 dose modifications/delays 3.3.1 lmb-100

LMB-100 infusion may be held for up to 72 hours due to drug-associated toxicity or adverse events from other intercurrent medical conditional (such as a primary cancer diagnosis) that resolve with or without medical intervention to grade 2 or less within this time frame. Toxicities for which further LMB-100 treatment should NOT be given or which are exceptions to the above guidelines are stated below in **Table 6**. The table also provides guidelines on how to manage some toxicities anticipated with LMB-100  
| Table 6. Guidelines for Managing Specific LMB-100 Adverse Events Event Action to Be Taken IRR/hypersensitivity reaction If an IRR/hypersensitivity develops, the infusion of LMB-100 should  be temporarily slowed down or interrupted. The patient should be  monitored until complete resolution of the symptoms and treated as  clinically indicated. Treatment or concomitant medication may  include acetaminophen, antihistamine, IV saline, oxygen,  bronchodilators, corticosteroids, and vasopressors depending on the  symptoms. If the infusion is interrupted: o In the event of IRR CTCAE Grade1, upon resolution of  symptoms, the infusion will resume at the same rate (the rate  being used at the time that the IRR occurred). o In the event of IRR Grade 2 or 3, upon resolution of symptoms,  the infusion will resume at one-half the previous rate. The  infusion can be re-escalated to initial rate if considered well  tolerated after 1 hour of infusion. o In the event of IRR CTCAE Grade 3, or CTCAE Grade 4  (which may include pulmonary or cardiac events) or an  anaphylactic reaction:  The infusion must be stopped and the patient should receive  aggressive treatment o Patients experiencing IRR CTCAE Grade 4 or anaphylaxis  must be permanently discontinued from LMB-100 treatment   |
|-----------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|  
| Event                                                                                                      | Action to Be Taken                                                                                                                                                                                                                                                                                                                                                                        |                                                          |
|------------------------------------------------------------------------------------------------------------|-------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|----------------------------------------------------------|
| Capillary leak syndrome                                                                                    | In the event of Grade ≥2 CTCAE capillary leak syndrome (urgent                                                                                                                                                                                                                                                                                                                            |                                                          |
| (CLS)                                                                                                      | intervention indicated): o Delay LMB-100 administration until complete resolution of the  event o For hypotension minimize fluid resuscitation to avoid fluid  overload Minimize crystalloid solutions (e.g., saline) o Vasopressor support (e.g., phenylephrine) if indicated to  stabilize blood pressure o Administer colloidal solutions (e.g., albumin) if there is a  clinically significant and persistent systolic blood pressure  drop, and the patient is symptomatic, or urine output declines o For pulmonary congestion provide diuretic and/or albumin  treatment in case of hypoalbuminemia as appropriate o Progressive shortness of breath may require in addition  endotracheal intubation or drainage of a pleural effusion o For oliguria and /or rising serum creatinine level delay LMB100 if Grade C3 urine output (<10 mL/hr) o Use fluids judiciously if increase in urine output is required o Use dopamine if patient is unresponsive to or unable to tolerate  fluids Monitor serum albumin levels prior to the LMB-100  treatment cycle o In the event of Grade ≥2 CTCAE pericardial effusion  (asymptomatic effusion small to moderate size), consider  delaying LMB-100 administration. In the event of Grade ≥3 CTCAE pericardial effusion (effusion with physiologic  consequences) stop LMB-100 treatment until full resolution                                                                                                                                                                                                                                                                                                                                                                                           |                                                          |
| Inflammatory reactions to                                                                                  | o                                                                                                                                                                                                                                                                                                                                                                                         | Hydrocortisone (200 mg IV) or equivalent dose of another |
| serosal membranes                                                                                          | corticosteroid as clinically indicated                                                                                                                                                                                                                                                                                                                                                    |                                                          |
| o                                                                                                          | In the event of pleuritis resulting in mild to severe pleuritic  pain, treat with analgesics or steroids as clinically indicated                                                                                                                                                                                                                                                          |                                                          |
| o                                                                                                          | For patients who have previously experienced pleuritis  consider administration of a tapering course of prednisone for  7 days starting with the next LMB-100 infusion                                                                                                                                                                                                                    |                                                          |
| Renal Toxicity                                                                                             | In the event of Grade 1 or greater renal toxicity consider increasing  oral or intravenous hydration, and consider delaying LMB-100  administration by up to 72 hours. In the event of Grade 2 or greater renal toxicity hold LMB-100  administration until recovery to Grade 1 or better. If this does not  occur within 72 hours, no further LMB-100 should be given during  the cycle. |                                                          |
| IRR = infusion related reaction; IV = intravenous; CTCAE = Common Terminology Criteria for Adverse  Events |                                                                                                                                                                                                                                                                                                                                                                                           |                                                          |
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3.3.2 pembrolizumab 3.3.2.1 dose delays

3.3.2.2 Cycle 3 day 1 or Cycle 2 day 1 (in the event that LMB-100 discontinued after cycle 1) of pembrolizumab may be delayed for up to 4 weeks if LMB-100 related toxicity has not resolved to grade 1 or better. If the delay exceeds 4 weeks, the patient will be removed from study therapy. Dose Modification and toxicity management for immune-related AEs associated with pembrolizumab AEs associated with pembrolizumab exposure may represent an immunologic etiology. These immune-related AEs (irAEs) may occur shortly after the first dose or several months after the last dose of pembrolizumab treatment and may affect more than on body system simultaneously. Therefore, early recognition and initiation of treatment is critical to reduce complications. Based on existing clinical study data, most irAEs were reversible and could be managed with interruptions of pembrolizumab, administration of corticosteroids and/or other supportive care. For suspected irAEs, ensure adequate evaluation to confirm etiology or exclude other causes. Additional procedures or tests such as bronchoscopy, endoscopy, skin biopsy may be included as part of the evaluation. Based on the severity of irAEs, withhold or permanently discontinue pembrolizumab and administer corticosteroids. Dose modification and toxicity management guidelines for irAEs associated with pembrolizumab are provided in **Table 7.** Table 7. Dose modification and toxicity management guidelines for immune related AEs associated with pembrolizumab General instructions:  
1. Corticosteroid taper should be initiated upon AE improving to Grade 1 or less and continue to taper over at least 4 weeks.  
2. For situations where pembrolizumab has been withheld, pembrolizumab can be resumed after AE has been reduced to Grade 1 or 0 and corticosteroid has been tapered. Pembrolizumab should be permanently discontinued if AE does not resolve within 12 weeks of last dose or corticosteroids cannot be reduced to ≤10 mg prednisone or equivalent per day within 12 weeks.  
3. For severe and life-threatening irAEs, IV corticosteroid should be initiated first followed by oral steroid. Other immunosuppressive treatment should be initiated if irAEs cannot be controlled by corticosteroids.  
| Immunerelated AEs               | Toxicity  grade or  conditions  (CTCAEv5.0)   | Action taken to       | irAE management   |                                                   |    |                                                 |
|---------------|-----------------------------------------------|-----------------------|-------------------|---------------------------------------------------|----|-------------------------------------------------|
| pembrolizumab | with corticosteroid  and/or other  therapies  | Monitor and follow-up |                   |                                                   |    |                                                 |
| Pneumonitis   | Grade 2                                       | Withhold              |                  | Administer  corticosteroids  (initial dose of 1-2 |   | Monitor participants for  signs and symptoms of |  
| Grade 3 or 4,  or recurrent  Grade 2                                                                | Permanently                                                                                                                                                                                                              | mg/kg prednisone                                                                                       |                                                                                                                                            |                                                                                                                                                                                                                   |                                                                                                                                       |
|-----------------------------------------------------------------------------------------------------|--------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|--------------------------------------------------------------------------------------------------------|--------------------------------------------------------------------------------------------------------------------------------------------|-------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|---------------------------------------------------------------------------------------------------------------------------------------|
| discontinue                                                                                         | or equivalent)  followed by taper                                                                                                                                                                                        | pneumonitis                                                                                            |                                                                                                                                            |                                                                                                                                                                                                                   |                                                                                                                                       |
|                                                                                                    | Evaluate participants with  suspected pneumonitis  with radiographic imaging  and initiate corticosteroid  treatment                                                                                                     |                                                                                                        |                                                                                                                                            |                                                                                                                                                                                                                   |                                                                                                                                       |
|                                                                                                    | Add prophylactic  antibiotics for  opportunistic infections                                                                                                                                                              |                                                                                                        |                                                                                                                                            |                                                                                                                                                                                                                   |                                                                                                                                       |
| Diarrhea /                                                                                          | Grade 2 or 3                                                                                                                                                                                                             | Withhold                                                                                               |                                                                                                                                            |                                                                                                                                                                                                                   |                                                                                                                                       |
| Colitis                                                                                             |                                                                                                                                                                                                                         | Administer  corticosteroids  (initial dose of 1-2  mg/kg prednisone  or equivalent)  followed by taper |                                                                                                                                           | Monitor participants for  signs and symptoms of  enterocolitis (i.e., diarrhea,  abdominal pain, blood or  mucus in stool with or  without fever) and of  bowel perforation  (i.e., peritoneal signs and  ileus). |                                                                                                                                       |
| Grade 4                                                                                             | Permanently  discontinue                                                                                                                                                                                                 |                                                                                                       | Participants with ≥ Grade  2 diarrhea suspecting  colitis should consider GI  consultation and  performing endoscopy to  rule out colitis. |                                                                                                                                                                                                                   |                                                                                                                                       |
|                                                                                                    | Participants with  diarrhea/colitis should be  advised to drink liberal  quantities of clear fluids.  If sufficient oral fluid  intake is not feasible, fluid  and electrolytes should be  substituted via IV  infusion. |                                                                                                        |                                                                                                                                            |                                                                                                                                                                                                                   |                                                                                                                                       |
| Aspartate  aminotransferase  / Alanine  aminotransferase  increased, or  blood bilirubin  increased | Grade 3 or 4                                                                                                                                                                                                             | Permanently                                                                                            |                                                                                                                                           | Administer                                                                                                                                                                                                        |                                                                                                                                       |
| discontinue                                                                                         | corticosteroids  (initial dose of 1-2  mg/kg prednisone  or equivalent)  followed by taper                                                                                                                               |                                                                                                        |                                                                                                                                            |                                                                                                                                                                                                                   |                                                                                                                                       |
| Grade 2                                                                                             | Withhold                                                                                                                                                                                                                 |                                                                                                       | Administer  corticosteroids  (initial dose of 0.5-  1 mg/kg prednisone  or equivalent)  followed by taper                                  |                                                                                                                                                                                                                  | Monitor with liver  function tests (consider  weekly or more frequently  until liver enzyme value  returned to baseline or is  stable |  
| Type 1 diabetes  mellitus  (T1DM) or  Hyperglycemia   | Newly onset  T1DM or  Grade 3 or 4  hyperglycemia  associated  with evidence  of -cell  failure   | Withhold                 |    | Initiate insulin  replacement  therapy for  participants with  T1DM                                    |    | Monitor participants for  hyperglycemia or other  signs and symptoms of  diabetes.                       |
|-------------------------------------------------------|----------------------------------------------------------------------------------------------------|--------------------------|-----|--------------------------------------------------------------------------------------------------------|-----|----------------------------------------------------------------------------------------------------------|
|                                                      | Administer antihyperglycemic in  participants with  hyperglycemia                                                                                                    |                          |     |                                                                                                        |     |                                                                                                          |
| Hypophysitis                                          | Grade 2                                                                                            | Withhold                 |    | Administer  corticosteroids and  initiate hormonal  replacements as  clinically  indicated.            |    | Monitor for signs and  symptoms of hypophysitis  (including  hypopituitarism and  adrenal insufficiency) |
| Grade 3 or 4                                          | Withhold or  permanently  discontinue1                                                             |                          |     |                                                                                                        |     |                                                                                                          |
| Hyperthyroidism                                       | Grade 2                                                                                            | Continue                 |    | Treat with nonselective betablockers (e.g.,  propranolol) or  thionamides as  appropriate                                                                                                        |    | Monitor for signs and  symptoms of thyroid  disorders.                                                   |
| Grade 3 or 4                                          | Withhold or  permanently  discontinue1                                                             |                          |     |                                                                                                        |     |                                                                                                          |
| Hypothyroidism                                        | Grade 2-4                                                                                          | Continue                 |    | Initiate thyroid  replacement  hormones (e.g.,  levothyroxine or  liothyroinine) per  standard of care |    | Monitor for signs and  symptoms of thyroid  disorders.                                                   |
| Nephritis and                                         | Grade 2                                                                                            | Withhold                 |     |                                                                                                        |     |                                                                                                          |
| Renal  dysfunction                                    | Grade 3 or 4                                                                                       | Permanently  discontinue |    | Administer  corticosteroids  (prednisone 1-2  mg/kg or  equivalent)  followed by taper.                |    | Monitor changes of renal  function                                                                       |
| Myocarditis                                           | Grade 1 or 2                                                                                       | Withhold                 |    | Based on severity  of AE administer  corticosteroids                                                   |    | Ensure adequate  evaluation to confirm  etiology and/or exclude  other causes                            |
| Grade 3 or 4                                          | Permanently  discontinue                                                                           |                          |     |                                                                                                        |     |                                                                                                          |
| All other  immune-related  AEs                        | Intolerable/  persistent  Grade 2                                                                  | Withhold                 |    | Based on type and  severity of AE  administer                                                          |    | Ensure adequate  evaluation to confirm  etiology and/or exclude                                          |  
| Grade 3                                                                                                              | Withhold or  discontinue  based on the  type of event.  Events that  require  discontinuation  include and not  limited to:  Guillain-Barré  Syndrome,  encephalitis   | corticosteroids   | other causes   |
|----------------------------------------------------------------------------------------------------------------------|------------------------------------------------------------------------------------------------------------------------------------------------------------------------|-------------------|----------------|
| Grade 4 or  recurrent  Grade 3                                                                                       | Permanently  discontinue                                                                                                                                               |                   |                |
| 1.                                                                                                                   | Withhold or permanently discontinue pembrolizumab is at the discretion of the investigator or treating  physician.                                                     |                   |                |
| NOTE:  For participants with Grade 3 or 4 immune-related endocrinopathy where withhold of pembrolizumab is required, |                                                                                                                                                                        |                   |                |  
1. Withhold or permanently discontinue pembrolizumab is at the discretion of the investigator or treating  
physician.  
NOTE: For participants with Grade 3 or 4 immune-related endocrinopathy where withhold of pembrolizumab is required, pembrolizumab may be resumed when AE resolves to ≤ Grade 2 and is controlled with hormonal replacement therapy or achieved metabolic control (in case of T1DM).  
3.3.2.3 Dose modification and toxicity management of infusion-reactions related to pembrolizumab Pembrolizumab may cause severe or life-threatening infusion-reactions including severe hypersensitivity or anaphylaxis. Signs and symptoms usually develop during or shortly after drug infusion and generally resolve completely within 24 hours of completion of infusion. Dose modification and toxicity management guidelines on pembrolizumab associated infusion reaction are provided in **Table 8**.  
| Table 8. Pembrolizumab Infusion Reaction Dose modification and Treatment Guidelines NCI CTCAE Grade Treatment Premedication at Subsequent  Dosing Grade 1 Increase monitoring of vital signs as medically  indicated until the participant is deemed medically  stable in the opinion of the investigator. None Mild reaction; infusion  interruption not  indicated; intervention  not indicated Grade 2 Stop Infusion. Participant may be  premedicated 1.5h (± 30  minutes) prior to infusion of  pembrolizumab with: Diphenhydramine 50 mg po  (or equivalent dose of  antihistamine). Acetaminophen 500-1000 mg  po (or equivalent dose of  analgesic).Requires therapy or  infusion interruption but  responds promptly to  symptomatic treatment  (e.g., antihistamines,  NSAIDs, narcotics, IV  fluids); prophylactic  medications indicated  for ≤24 hrs Additional appropriate medical therapy may  include but is not limited to: IV fluids Antihistamines NSAIDs Acetaminophen Narcotics   |
|--------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|  
| NCI CTCAE Grade                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                           | Treatment                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                           | Premedication at Subsequent  Dosing   |
|-----------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|---------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|---------------------------------------|
| Increase monitoring of vital signs as medically  indicated until the participant is deemed medically  stable in the opinion of the investigator. If symptoms resolve within 1 hour of stopping drug  infusion, the infusion may be restarted at 50% of  the original infusion rate (e.g. from 100 mL/hr to 50  mL/hr). Otherwise dosing will be held until  symptoms resolve and the participant should be  premedicated for the next scheduled dose. Participants who develop Grade 2 toxicity  despite adequate premedication should be  permanently discontinued from further study  drug treatment                                                                                                    |                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                     |                                       |
| Grades 3 or 4                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                             | Stop Infusion.                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                      | No subsequent dosing                  |
| Grade 3:                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                  | Additional appropriate medical therapy may  include but is not limited to: Epinephrine** IV fluids Antihistamines NSAIDs Acetaminophen Narcotics Oxygen Pressors Corticosteroids Increase monitoring of vital signs as medically  indicated until the participant is deemed medically  stable in the opinion of the investigator. Hospitalization may be indicated. **In cases of anaphylaxis, epinephrine should be  used immediately. Participant is permanently discontinued from  further study drug treatment. |                                       |
| Prolonged (i.e., not  rapidly responsive to  symptomatic medication  and/or brief interruption  of infusion); recurrence  of symptoms following  initial improvement;  hospitalization indicated  for other clinical  sequelae (e.g., renal  impairment, pulmonary  infiltrates) Grade 4: Life-threatening; pressor  or ventilatory support  indicated Appropriate resuscitation equipment should be available at the bedside and a physician readily available during the period of  drug administration. For further information, please refer to the Common Terminology Criteria for Adverse Events v5.0 (CTCAE) at  http://ctep.cancer.gov 3.3.2.4 Other allowed dose interruptions for pembrolizumab |                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                                     |                                       |  
Pembrolizumab may be interrupted for situations other than treatment-related AEs such as medical or surgical events or logistical reasons not related to study therapy. Participants should be placed back on study therapy within 3 weeks of the scheduled interruption. The reason for interruption should be documented in the patient's study record.
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3.4 study calendar 1 cycle

Screening assessments will occur within 28 days prior to enrollment unless otherwise indicated. If screening assessments are performed within 3 days prior to enrollment, the assessments do not need to be repeated on C1D1 unless otherwise indicated. Assessments after C1D1 may be performed up to 3 days prior to indicated time unless otherwise indicated. LMB-100 dosing cycles after cycle 1 may be delayed for up to two weeks to accommodate schedule conflicts, federal holidays and inclement weather, etc. Pembrolizumab dosing cycles may be delayed up to three weeks for the same reasons, however, cycle 3 day 1 (or Cycle 2 day 1 in the event that LMB-100 discontinued after cycle 1) may be delayed for longer as described in section **3.3.2**.  
| event that LMB-100 discontinued after cycle 1) may be delayed for longer as described in section 3.3.2. LMB-100 Cycle(s)  Subsequent Cycles (starting  at cycle 2 or cycle 3)  (cycle 1 +/- cycle 2)  (for a maximum of 2 years  after initial pembrolizumab  therapy and, if applicable,  through second course)14  (1 cycle=21days1 )   | Safety FollowUp Visit (_~30  days and ~90  days after  completion of  study therapy)15      | Long-Term   |     |     |     |           |       |    |
|-------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|------|-------------|-----|-----|-----|-----------|-------|----|
| Follow Up (every  6 - 12 weeks)16                                                                                                                                                                                                                                                                                                         |      |             |     |     |     |           |       |    |
| Screening                                                                                                                                                                                                                                                                                                                                           | Day  | Day         | Day | Day | Day |           |       |    |
| Procedure                                                                                                                                                                                                                                                                                                                                 | 1    | 3           | 4   | 5   | 8   | Day  1513 | Day 1 |    |
| LMB-100                                                                                                                                                                                                                                                                                                                                   | X    | X           | X   |     |     |           |       |    |
| Pembrolizumab                                                                                                                                                                                                                                                                                                                             | X    |             |     |     |     |           |       |    |
| History and PE                                                                                                                                                                                                                                                                                                                            | X    | X           | X   | X   | X   | X         | X     | X  |
| Weight                                                                                                                                                                                                                                                                                                                                    | X    | X           | X   | X   | X   | X         | X     |    |
| Height                                                                                                                                                                                                                                                                                                                                    | X    |             |     |     |     |           |       |    |
| Vital signs2                                                                                                                                                                                                                                                                                                                              | X    | X           | X   | X   | X   | X         | X     | X  |
| Performance Score                                                                                                                                                                                                                                                                                                                         | X 16 | X           | X   |     |     |           |       |    |
| Labs3, 18                                                                                                                                                                                                                                                                                                                                 | X 4  | X 5         | X   | X   | X   | X         | X     | X  |
| TSH, free T4, total T3                                                                                                                                                                                                                                                                                                                    | X    | X           | X   |     |     |           |       |    |
| Urinalysis                                                                                                                                                                                                                                                                                                                                | X    | X           | X   |     |     |           |       |    |  
| LMB-100 Cycle(s)                                          | Subsequent Cycles (starting  at cycle 2 or cycle 3)                                                           |                               |                        |     |     |           |       |
|-----------------------------------------------------------|---------------------------------------------------------------------------------------------------------------|-------------------------------|------------------------|-----|-----|-----------|-------|
| (cycle 1 +/- cycle 2)                                     | (for a maximum of 2 years  after initial pembrolizumab  therapy and, if applicable,  through second course)14 |                               |                        |     |     |           |       |
| (1 cycle=21days1 )                                        | Safety FollowUp Visit (_~30  days and ~90  days after  completion of  study therapy)15                                                                                                               | Long-Term                     |                        |     |     |           |       |
| Follow Up (every  6 - 12 weeks)16                         |                                                                                                               |                               |                        |     |     |           |       |
| Screening                                                           | Day                                                                                                           | Day                           | Day                    | Day | Day |           |       |
| Procedure                                                 | 1                                                                                                             | 3                             | 4                      | 5   | 8   | Day  1513 | Day 1 |
| HLA Typing (Class                                         | 6                                                                                                             |                               |                        |     |     |           |       |
| I and Class II)                                           | X                                                                                                             |                               |                        |     |     |           |       |
| Urine or serum hCG  in women of  childbearing  potential7 | X 8                                                                                                           | X 8                           | X                      | X   |     |           |       |
| Confirmation of dx9                                       | X                                                                                                             |                               |                        |     |     |           |       |
| NIH Advance                                               | 10                                                                                                            |                               |                        |     |     |           |       |
| Directives Form                                           | X 6                                                                                                           | Post-LMB-100 and after second |                        |     |     |           |       |
| Biopsy (optional)                                         | X17                                                                                                           | X                             | cycle of pembrolizumab |     |     |           |       |
| Correlative                                               | See section 5.2                                                                                               |                               |                        |     |     |           |       |
| Research Studies  CT CAP and/or                           | Every 6 weeks ± 7 days                                                                                        | X                             |                        |     |     |           |       |
| MRI                                                       | X                                                                                                             | 11                            | X                      |     |     |           |       |
| FDG CT/-PET                                               | X                                                                                                             | Every 6 weeks ± 7 days        | X 11                   | X   |     |           |       |
| ECG12                                                     | X                                                                                                             | X                             | X                      | X   | X   | X         |       |
| Echocardiogram                                            | X                                                                                                             |                               |                        |     |     |           |       |
| Adverse Events                                            | Monitored continuously                                                                                        |                               |                        |     |     |           |       |  
| LMB-100 Cycle(s)                                   | Subsequent Cycles (starting  at cycle 2 or cycle 3)                                                                                                                        |           |     |     |     |           |       |
|----------------------------------------------------|----------------------------------------------------------------------------------------------------------------------------------------------------------------------------|-----------|-----|-----|-----|-----------|-------|
| (cycle 1 +/- cycle 2)                              | (for a maximum of 2 years  after initial pembrolizumab  therapy and, if applicable,  through second course)14                                                              |           |     |     |     |           |       |
| (1 cycle=21days1 )                                 | Safety FollowUp Visit (_~30  days and ~90  days after  completion of  study therapy)15                                                                                                                                                                            | Long-Term |     |     |     |           |       |
| Follow Up (every  6 - 12 weeks)16                  |                                                                                                                                                                            |           |     |     |     |           |       |
| Screening                                                    | Day                                                                                                                                                                        | Day       | Day | Day | Day |           |       |
| Procedure                                          | 1                                                                                                                                                                          | 3         | 4   | 5   | 8   | Day  1513 | Day 1 |
| Concomitant                                        | Monitored continuously                                                                                                                                                     |           |     |     |     |           |       |
| Medications Telephone/email  follow-up q 3  months | X                                                                                                                                                                          |           |     |     |     |           |       |
| 1                                                  | Most subjects are expected to complete 2 cycles of LMB-100; however, in the case of RECIST or clinical disease progression, LMB-100 may be discontinued after 1 cycle with |           |     |     |     |           |       |  
1 Most subjects are expected to complete 2 cycles of LMB-100; however, in the case of RECIST or clinical disease progression, LMB-100 may be discontinued after 1 cycle with pembrolizumab initiated in cycle 2 instead.  
2 **At screening:** heart rate, blood pressure, body temperature, pulse oximetry. **During the infusions** of LMB-100 in Cycle 1, vital signs (including, if possible, supine diastolic and systolic blood pressure, pulse rate, and temperature) must be monitored pre-infusion and every 15 minutes (± 5 minutes) until the end of the infusion. Following the completion of the infusion, vital signs should be checked every 30 minutes (± 10 minutes) for the first 2 hours and once at 4 hours after infusion completion. On days when there is no infusion, vital signs should be monitored per standard of care.  
3 CBC with differential, Acute Care Panel (Na, K, Cl, CO2, creatinine, glucose, BUN), Hepatic Panel (ALP, AST, ALT, total & direct bilirubin), Mineral Panel (albumin, Ca, Mg, P), lactate dehydrogenase, creatine kinase, C-reactive protein, PT, PTT, fibrin degradation products.  
4 Performed within 10 days prior to start of study therapy.  
5 Only required if more than 10 days have passed since screening assessment. If eligibility criteria are not met at this timepoint, subject may not be treated. 6 May be performed after study consent is signed but prior to treatment initiation (baseline). 7 Required in women of childbearing potential (see **Appendix B** for definition). 8 Performed within 3 days prior to enrollment. Must be repeated on C1D1 (pre-treatment) if more than 72 hours have passed since screening assessment and actual treatment initiation.  
9 This includes confirmation of histology, mutation status as required and mesothelin expression. Please see section 2.2 for tissue requirements 10 As indicated in section **11.3**, all subjects will be offered the opportunity to complete an NIH advance directive form. This should be done preferably at baseline but can be done at any time during the study as long as the capacity to do so is retained. The completion of the form is strongly recommended but is not required.  
11 PET is optional at this timepoint and may be performed if feasible.  
12 Single 12-lead ECG will be recorded at screening, then pre- and end of LMB-100 infusion for first cycle and at the withdrawal and follow-up visit. Pre-infusion at all other study drug administrations. Additional unscheduled ECG assessments should be performed if cardiovascular symptoms or abnormalities occur.  
13 Day 15 laboratory assessments should be performed -3/+1 day and may be performed outside of NIH. History may be performed remotely, and physical exam is only required in subjects seen at NIH.  
14 Per section **3.1.2**, some subjects may be eligible for up to an additional 17 cycles of therapy, called the second course. Surveillance during the second course will be the same as during the initial course unless otherwise indicated. See **Appendix D** for further details on second course imaging.  
15 Follow up visit will occur within +/- 1 week of indicated time (4- 6 weeks **after the last dose of study drug (first course and second course)**. The assessments listed refer to those that will be performed if the patient is seen in clinic **(first course and second course)**. If the patient is unable to return to the clinic for the follow up visit, adverse event and ECOG assessments will be performed by telephone or email, we will request that required labs, scans and ECG evaluations be performed locally and provided to us, and if the patient has visited a local oncologist in this timeframe, the progress notes will also be requested.. Scans performed only in patients who have not had progressive disease on pembrolizumab. Scans will continue every 6 weeks until disease progression or start of a new anti-cancer treatment. After disease progression on pembrolizumab or in those subjects who were removed from study therapy for reasons other than disease progression, subjects will be followed every 12 weeks by telephone/email for assessment of survival status, adverse events and initiation of new anti-cancer therapy.  
16 Performed within 7 days prior to start of study therapy.  
17 Performed if no archival tumor tissue is available for diagnosis confirmation. Tissue samples will also be sent to Mr. Liqiang Xi in the Laboratory of Pathology for biomarker analysis.  
18 In addition to the labs noted in the Study Calendar, clinical labs (mineral panel, hepatic panel, CBC with differential, CRP, UA) are to be drawn daily while patients are admitted for LMB-100 administration.
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3.5 cost and compensation 3.5.1

NIH does not bill health insurance companies or participants for any research or related clinical care that participants receive at the NIH Clinical Center. If some tests and procedures performed outside the NIH Clinical Center, participants may have to pay for these costs if they are not covered by insurance company. Medicines that are not part of the study treatment will not generally be provided or paid for by the NIH Clinical Center. 3.5.2 Compensation Participants will not be compensated on this study. 3.5.3 Reimbursement The NCI will cover the costs of some expenses associated with protocol participation. Some of these costs may be paid directly by the NIH and some may be reimbursed to the participant/guardian as appropriate. The amount and form of these payments are determined by the NCI Travel and Lodging Reimbursement Policy.
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3.6 criteria for removal from

Prior to removal from study, effort must be made to have all subjects complete a safety visit approximately 90 days following the last dose of study therapy. 3.6.1 Criteria for removal from protocol therapy  
Completion of protocol therapy o Note: The number of treatments is calculated starting with the first dose of Pembrolizumab. Participants who stop Pembrolizumab after receiving 35 doses may be eligible for retreatment if they progress after stopping study treatment provided they meet the requirements detailed in section 3.1. Participants may be retreated in the Second Course Phase (Retreatment) for up to an additional 17 cycles (approximately 1 year).  
Progressive disease on pembrolizumab Noncompliance with study treatment or procedure requirements
Requirement for use of prohibited medications/vaccines (see **4.1.2**)  
Recurrent Grade 2 pneumonitis Participant (or LAR if one was designated when participant become incapacitated during study) requests to be withdrawn from active therapy Unacceptable Toxicity as defined in section 3.3 Investigator discretion Positive pregnancy test 3.6.2 Off-Study Criteria
Investigator decision to end the study Participant requests to be withdrawn from study  
Lost to follow up Death Screen failure 3.6.3 Lost to Follow-up
A participant will be considered lost to follow-up if he or she fails to return for 3 scheduled visits and is unable to be contacted by the study site staff. The following actions must be taken if a participant fails to return to the clinic for a required study visit:  
The site will attempt to contact the participant and reschedule the missed visit within 3 business days and counsel the participant on the importance of maintaining the assigned visit schedule and ascertain if the participant wishes to and/or should continue in the study.  
Before a participant is deemed lost to follow-up, the investigator or designee will make every effort to regain contact with the participant (where possible, 3 telephone calls and, if necessary, an IRB approved certified letter to the participant's last known mailing address or local equivalent methods). These contact attempts should be documented in the participant's medical record or study file.  
Should the participant continue to be unreachable, he or she will be considered to have withdrawn from the study with a primary reason of lost to follow-up.
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4 concomitant medications/measures

All concomitant medications received within 28 days before the first dose of trial treatment and 90 days after the last dose of trial treatment (first course and if applicable, second course) should be recorded. Concomitant medications administered after 90 days after the last dose of trial treatment (first course and if applicable, second course) should be recorded for SAEs and ECIs as defined in section 7.2.
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4.1.1 permitted therapy

All treatments that the investigator considers necessary for a participant's welfare may be administered at the discretion of the investigator in keeping with the community standards of medical care. All concomitant medication will be recorded on the case report form (CRF) including all prescription, over-the-counter (OTC), herbal supplements, and IV medications and fluids. If changes occur during the trial period, documentation of drug dosage, frequency, route, and date may also be included on the CRF. Supportive care may be administered according to NIH CC Pharmacy Guidelines unless otherwise specified in section 3.3.
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4.1.2 prohibited therapy

Patients should be treated for all concomitant conditions and adverse events according to accepted standards of medical care at the discretion of the investigator. The following treatments are not permitted while patient is receiving study therapy:  
Antineoplastic systemic chemotherapy or biological therapy Immunotherapy not specified in this protocol Chemotherapy not specified in this protocol Investigational agents other than the study agents Radiotherapy. Note: Radiation therapy to a symptomatic solitary lesion or to the brain may be allowed at the investigator's discretion.  
Live vaccines. Examples of live vaccines include, but are not limited to, the following:
measles, mumps, rubella, varicella/zoster, yellow fever, rabies, BCG, and typhoid vaccine. Seasonal influenza vaccines for injection are generally killed virus vaccines and are allowed; however, intranasal influenza vaccines (e.g., FluMist®) are live attenuated vaccines and are not allowed.  
Systemic glucocorticoids for any purpose other than to modulate symptoms from an event of clinical interest of suspected immunologic etiology. The use of physiologic doses of corticosteroids may be considered per investigator discretion.  
Strong inhibitors or inducers of CYP3A4 (as can be found here: Flockhart Table or in another frequently updated source)
Participants who, in the assessment by the investigator, require the use of any of the aforementioned treatments for clinical management should be removed from the study. All treatments that the Investigator considers necessary for a participant's welfare may be administered at the discretion of the Investigator in keeping with the community standards of medical care. Medications or vaccinations specifically prohibited in the exclusion criteria are not allowed during the ongoing study. If there is a clinical indication for any medication or vaccination specifically prohibited during the study, discontinuation from study therapy is required. The final decision on any supportive therapy or vaccination rests with the investigator and/or the participant's primary physician. There are no prohibited therapies during the Post-Treatment Follow-up Phase.
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5 correlative studies for research

All blood samples for PK assessment will be collected from an IV line different to that receiving the infusion to measure free and total concentrations for LMB-100 for all patients. The date and time of each sample collection will be recorded. If multiple samples are drawn at a given time point, the PK sample should take precedence. Free and total plasma concentrations of LMB-100 will be measured using validated ligand-binding assays. 5.1.1.1 Sample collection: Blood for PK samples should be drawn from the opposite extremity as the one used for drug delivery. If feasible, a peripheral line used for this purpose should remain in place until the last scheduled collection for a given day. Blood will be collected in 2 mL K2EDTA tubes (purple top)
at the times defined in section 5.2. Samples should be inverted 8 to 10 times after collection. Store on wet ice or at 4°C. Processing within 60 minutes of blood collection is highly preferred. 5.1.1.2 Sample processing Samples will be processed in the Clinical Pharmacology Program. Please e-mail NCIBloodcore@mail.nih.gov at least 24 hours before transporting samples (the Friday before is preferred). For sample pickup, page 102-11964. For immediate help, call 240-760-6180 (main blood processing core number) or, if no answer, 240-760-6190 (main clinical pharmacology lab number). For questions regarding sample processing, contact at NCIBloodcore@mail.nih.gov by e-mail or at 240-858-3191. Upon arrival in the CPP the following procedures should be followed:
1. Store on wet ice until centrifugation.  
2. Centrifuge 1500xg for 10 minutes at 4°C within 60 minutes of blood collection. 3. Transfer plasma specimen to 2mL cryovials and store at -70°C.
The analyses will be performed retrospectively in batched samples or at the end of the trial.  
5.1.1.3 Sample Shipping
Samples will be shipped by the CPP on dry ice to Ms. Yanyu Wang in Frederick for analysis.  
Leidos Biomedical Research, Inc. Attention: Ms. Yanyu Wang & Dr. Jon Inglefield Building 469, Room 120 Miller Drive Frederick, MD 21702 Phone: 301-846-6905/301-846-6865 5.1.1.4 Sample storage Samples will be stored in the CPP until shipment to the Leidos Biomedical Inc. Lab in Frederick. 5.1.2 Assessment of anti-drug antibodies (ADAs)  
5.1.2.1 Sample Collection
Samples will be per the schedule in section **5.2.**
Draw 2mL blood into K2EDTA tube (purple top). Samples should be inverted 8 to 10 times after collection. Store on wet ice or at 4°C. Processing within 60 minutes of blood collection is highly preferred. 5.1.2.2 Sample Processing Samples will be processed in the Clinical Pharmacology Program. Please e-mail:NCIBloodcore@mail.nih.gov at least 24 hours before transporting samples (the Friday before is preferred). For sample pickup, page 102-11964. For immediate help, call 240-760-6180 (main blood processing core number) or, if no answer, 240-760-6190 (main clinical pharmacology lab number). For questions regarding sample processing, contact NCIBloodcore@mail.nih.gov . Upon arrival in the CPP, each sample should be processed in the following manner:
1. Store on wet ice until centrifugation.  
2. Centrifuge 1500xg for 10 minutes at 4°C within 60 minutes of blood collection. 3. Transfer plasma specimen to 2mL cryovials and store at -70°C.
Autoantibody levels will be retrospectively assessed. 5.1.2.3 Sample Shipping Samples will be shipped by the CPP on dry ice to Ms. Yanyu Wang in Frederick for analysis.  
Leidos Biomedical Research, Inc. Attention: Ms. Yanyu Wang, Dr. Jon Inglefield Building 469, Room 120 Miller Drive Frederick, MD 21702 Phone: 301-846-6905/301-846-6865 5.1.2.4 Sample Storage Samples will be stored in the CPP until shipment to the Leidos Biomedical Inc. Lab in Frederick. 5.1.3 Retrospective Analysis of Mesothelin, KRAS and PD-L1 Expression in tumor tissue The mesothelin expression analysis and KRAS mutation status will be performed by the NCI Lab of Pathology at the time of histological confirmation of non-squamous non-small cell lung cancer. This must be done prior to subject enrollment to trial and treatment. Mesothelin expression can be analyzed on archival tumor tissue sample or newly obtained fresh core or excisional biopsy of a tumor lesion not previously irradiated. Leftover tissue from archival specimens or tumor biopsies obtained at baseline or from optional collections at postLMB-100 (either post cycle 1 or post cycle 2 depending upon progression), and post second cycle of pembrolizumab may be used for this purpose. Specimens will be used to correlate treatment response with mesothelin expression and with anti PD-L1 in exploratory analyses. Additionally, this tissue will undergo IHC staining to analyze CD4 and CD8 T-Cell subsets. 5.1.3.1 Specimen collection Collection of optional tumor biopsies should be guided by ultrasound, CT scan, or other method according to the location of the selected lesion using a ≤ 18-gauge needle to provide cores ideally of at least 20 mm in length or equivalent size. At least 3, ideally 4 core biopsies will be obtained at each time point (baseline, post cycle 2, and post pembrolizumab). Fine needle aspiration and biopsy of bone lesions are not acceptable. All biopsies collected under this protocol will undergo review in the NCI Laboratory of Pathology. 5.1.4 Mesothelin and Megakaryocyte Potentiating Factor (MPF) Serum Samples The levels of serum mesothelin as well as megakaryocyte potentiating factor, which is released into serum from the processing of mesothelin precursor protein will be assessed in order to determine correlation with therapeutic response.
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5.1.4.1 sample collection

Samples will be obtained prior to the first LMB-100 dose in cycle 1 and at the end of treatment All blood samples will be taken by either direct venipuncture or an indwelling venous access. At each sample collection time, blood (2mL) will be drawn in a 3.5 mL (red) serum separator tube labeled as follows:
- Subject ID Number - Study Number - Time and date of collection  
5.1.4.2 Sample Processing
Please e-mail NCIBloodcore@mail.nih.gov at least 24 hours before transporting samples (the Friday before is preferred). For sample pickup, page 102-11964. For immediate help, call 240-760-6180 (main blood processing core number) or, if no answer, 240-760-6190 (main clinical pharmacology lab number). For questions regarding sample processing, contact NCIBloodcore@mail.nih.gov. Upon arrival in the CPP, each sample should be processed in the following manner: Allow blood to clot for 10 minutes and centrifuge to separate the serum within 30 minutes of collection. If unable to process within 30 minutes, then whole blood tubes may be stored upright in refrigerator (4-8C) for up to 48 hours prior to processing. Processing of samples within 30 minutes is strongly preferred. Stability studies will establish if degradation of soluble mesothelin in whole blood during 0.5 to 48 hours is significant and therefore if the data from these samples should be included in the analysis. Transfer the serum into two pre-labeled cryotubes and immediately freeze by placing on dry ice.  
Transfer frozen serum samples into a - 80C freezer for storage.  
5.1.4.3 Sample Storage All serum samples will be stored by Dr. Figg's Clinical Pharmacology Program.  
5.1.5 Gene expression-based characterization of the immune landscape before and after treatment with LMB-100 and pembrolizumab
As referenced in section **5.1.3.1**, optional tumor biopsies will be performed in consenting patients when deemed feasible at the following time points: before initiation of therapy at baseline (archival tissue may also be used), at the end of cycle 2, and after pembrolizumab. We will evaluate tumor biopsies before and after treatment with LMB-100 and after treatment with pembrolizumab using a hybridization-based digital gene expression platform nCounter (NanoString Technologies). This platform allows for unbiased multiplexed quantification of RNA transcripts achieving sensitivity comparable to quantitative reverse-transcription polymerase chain reaction (Q-RT-PCR), without any enzymatic reaction involved in the process. We have expertise in the use of this technology for the characterization of purified cell subsets and for the study of the changes that occur in human tumors, as a consequence of a given treatment, using tumor core biopsy tissues. This analysis can be performed on flash frozen biopsies without any further isolation or enrichment of specific cell types. For the present study, we will screen pre- and post-treatment samples for the expression of markers of immune cell subsets (CD3, CD8, CD4, etc.), local production of cytokines (interferon-gamma, tumor necrosis factor-alpha, etc.) and chemokines (CXCL13, CCL5, etc.), adhesion molecules and others. The purpose of this study is to characterize the molecular changes that occur within the tumors following treatment with LMB-100 and pembrolizumab. The ultimate goal is to gain a better understanding of the mechanism of action of this treatment, and to identify molecular correlates of clinical outcomes such as objective responses and/or improved survival. In order to achieve these goals, flash frozen samples will be subjected to total RNA isolation followed by hybridization with capture and detection probes specific for 620 transcripts, including genes involved in the regulation of the immune function and markers expressed by tumor cells and tumor stroma. To cover those target transcripts, a combination of a commercially available predesigned probe set (GX Human Immunology v2, NanoString Technologies) will be used in combination with a custom-designed code set of thirty additional targets (Panel Plus, NanoString Technologies). Hybridization complexes will be quantified using a NanoString nCounter Analysis System, at the Genomics Core Facility of the Center for Cancer Research, NCI. Results obtained by this approach will be correlated with data obtained from immunohistochemistry of tumor biopsies as well as with data from analysis of peripheral blood populations, for a comprehensive study of the mechanism of action of LMB-100 in mesothelioma patients. Samples will be stored in the Laboratory of Dr. Raffit Hassan, Building 10, Room 3B51.
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5.1.6 biomarker analysis

If not previously completed, every patient entering the trial will have tumor tissue analyzed with the the TruSight Oncology 500 Assay in the Molecular Diagnostics Section of the Laboratory of Pathology. If previously completed, the existing results will be used to inform study results. This is a clinical next-generation sequencing-based multiple-biomarker assays that targets the most relevant single nucleotide variants, small insertions and deletions, copy number variants, and gene fusions from 161 cancer genes. Tissue samples will be sent to Mr. Liqiang Xi at LP for analysis. Phone: 301-480-8933 Email: xil2@mail.nih.gov 5.1.7 Circulating tumor DNA (ctDNA) before and sequentially after treatment with LMB-100 and pembrolizumab:
For post-treatment correlative study, based on comprehensive data analysis (as explained above) we will select 1-3 mutation targets per patient as circulating tumor DNA (ctDNA) markers to be used for monitoring the treatment response. The customized digital droplet PCR (ddPCR) mutation detection assay will be performed on a QX200 ddPCR system (Bio-Rad Laboratories, Inc) with 10-20 ng cell-free DNA extracted from about 4 mL plasma for each time point. Two time points before treatment and as many as time points during/after treatment will analyzed. Specifically, blood samples will be collected at baseline, pre/post LMB-100 on days 1, 3, 5 followed by day 8, 15 (-3/+1 day of C1D15 and C2D15) for each cycle and end of treatment using standard procedures and transferred to Figg's lab on a patient by patient basis. Plasma preparation /storage will be done at Figg lab as described below. Once the plasma samples are collected for few patients in a batch, PI will request to Laboratory of Pathology for ddPCR mutation detection assay. Samples will be sent by the Figg lab on dry ice to Mr. Liqiang Xi at LP for analysis. Phone: 301-480-8933 Email: xil2@mail.nih.gov There will also be blood drawn pre/post pembrolizumab on each subsequent cycle. The ultimate goal is to identify ctDNA correlates of clinical outcomes such as objective responses and/or improved survival to LMB-100 with pembrolizumab. Plasma preparation for cell free (cf)DNA extraction using Streck collection tubes: Streck tubes can be stored for processing when Figg's lab is open. Collect 2- 10 ml brown/black Streck cell free DNA/ cf DNA (brown black tiger top). Samples are stable for up to 5 days at room temperature.  
1. In hood: transfer blood to 15 mL conical tube 2. Centrifuge 15 mL conical tubes for 10 minutes at 1500 ± 150xg (=1500RCF) 4° 3. Transfer supernatant to a fresh 15 ml tube without disturbing the leukocyte layer 4. Centrifuge plasma a second time for 10 min at 3000 ± 150xg (=3000 RCF) 4° 5. Transfer supernatant to a fresh 15 ml centrifuge tube without disturbing the cellular layer.  
Leave a residual volume of about 0.3 ml (~7 mm) on the bottom of the 15 ml tube to avoid cellular contamination 6. After transferring the plasma to a new 15 ml centrifuge tube as described, gently mix plasma and record total plasma volume (typically ~ 4 ml plasma per 10 ml blood)
7. Make 1 mL aliquots into standard cryovials 8. Store plasma in freezer at -80°C
5.1.8 Pleural effusion to establish cell lines and cytology studies
In the case that patients present with or develops a pleural effusion who are symptomatic may proceed with therapeutic thoracentesis during any time point during the course of treatment at the IR suite or at bedside from a pigtail catheter. Fluid that is collected will be analyzed and used for research purposes including cytology and establishing cell-lines. All samples will be coded. 5.1.9 Cytokines and circulating endothelial cells (CECs) for identification of a mechanism for Pseudomonas exotoxin A (PE)-mediated capillary leak syndrome (CLS)
PE-based RITs cause dose-limiting CLS. At low doses CLS manifests as mild and transient weight gain, hypoalbuminemia, and peripheral or facial edema. At higher doses it can cause lifethreatening cardiopulmonary compromise. Previous studies in rats have indicated that pathological changes indicative of CLS onset occur within just two hours of toxin administration and even when the PE fragment lacks a targeting domain (40). *In vitro* studies with cultured endothelial cells have demonstrated that super-physiologic doses of PE-based RITs cannot induce endothelial cell toxicity unless the cells express the RIT target (41). Together these data suggest the hypothesis that PE-based RITs cause CLS by triggering release of vasoactive cytokines by specific immune cells rather than through direct damage to endothelial cells. To test this hypothesis, we will collect additional blood from participants. Levels of a panel of cytokines known to affect the vasculature will be assessed. Change in cytokine levels from baseline will be compared. In addition, circulating endothelial progenitor cells (CEP) and mature circulating endothelial cells (CEC) will be collected to assess endothelial cell dynamics (see Section 5.2 for specific time points). These cells will be assessed by multiparameter flow cytometry. Cells will be analyzed for forward and side scatter, and cells expressing hematopoietic markers will be excluded. Endothelial cells will be identified using co-expression of markers, such as CD31 and CD146 for CEC, and CD31 and CD133 for CEP. The cell populations will also be analyzed for viability using scatter profiles and a vital stain, such as Hoechst 33258. Percentages of stained cells will be determined and compared with appropriate negative controls. Multiparameter flow cytometric analysis will be performed with a Miltenyi Quant equipped with FlowJo software, using a minimum of 100,000 events per analysis. 5.1.9.1 Specimen collection- cytokine analysis Two mL of plasma and serum will be collected in SST (red top) tubes from participants on the days and time points shown in Sections 5.2. 5.1.9.2 Sample processing- cytokine analysis Samples will be processed in the Clinical Pharmacology Program as described in Section **5.1.1.2.** Upon arrival in the CPP, each plasma sample should be processed in the following manner:  
1. Store on wet ice until centrifugation. 2. Centrifuge 1500xg for 10 minutes at 4°C within 60 minutes of blood collection. 3. Immediately transfer plasma specimen (maintaining samples at 4°C throughout handling process) to 2mL cryovials and store at -70°C.  
Upon arrival in the CPP, each serum sample should be processed in the following manner:
1. Store at room temperature for 15-30 minutes to allow blood to clot  
2. Centrifuge at 1500xg for 10 minutes at 4°C to remove clot
3. Immediately transfer serum specimen (maintaining samples at 4°C throughout handling process) to 2mL cryovials and store at -70°C.  
5.1.9.3 Sample Storage- cytokine analysis Samples will be stored in the CPP. Deidentified samples will be transferred to TGMB or NCIFrederick for analysis within 12 months of freezing.  
5.1.9.4 Sample Collection- CECs Draw blood into one 8-cc CPT citrate (BD) tube as specified in Section 5.2.
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5.1.9.5 sample processing-cecs

Contact the Trepel Lab, Developmental Therapeutics Branch, NCI by email (Jane Trepeltrepel@helix.nih.gov and Sunmin Lee- lees@pop.nci.nih.gov) when the patient is scheduled and by phone as soon as the blood is drawn at 240-760-6330 and a lab member will pick up the sample.  
| 5.2                                                                         | SAMPLE COLLECTION SCHEDULE    |                                 |                                                    |                                            |                                |
|-----------------------------------------------------------------------------|-------------------------------|---------------------------------|----------------------------------------------------|--------------------------------------------|--------------------------------|
| Cycle                                                                       | Daye                          | PK                              | ADA                                                | Tumor  Sampl e                             | Serum                          |
| mesothelin  and MPF                                                         | ctDNA  Blood  sample          | Pleural  effusion               | Cytokines  & CECs                                  |                                            |                                |
| (5.1.1)                                                                     | (5.1.2)                       | (5.1.8)                         | (5.1.9)                                            |                                            |                                |
| (5.1.3,  5.1.5,  5.1.6)                                                     | (5.1.4)                       | (5.1.7)                         |                                                    |                                            |                                |
| 2 mL blood  in K2EDTA  tube (purple  top)                                   | 2 mL                          |                                 |                                                    |                                            |                                |
| blood in  K2EDTA  tube  (purple  top)                                       | N/A                           | 2 mL blood  in 3.5 mL  SST tube | 10 mL  cfDNA in  brown /  black  streck  tiger top | 1 Liter  (vacuum  drainage  container)     | 2 mL SST  tube, 8 mL CPT  tube |
| Screening                                                                   | Screeni ng  period  ± 3  days | Xa                              | Screening                                          | Any time  during the  course of  treatment |                                |
| 1                                                                           | 1                             | Pre-dose,                       |                                                    |                                            |                                |
| EOI, 1, 2, 3,  4, and 6  hours after  start of  infusion                    | Pre-dose                      | Xb                              | X                                                  | Pre                                            |                                |
| dose/EOI  on days 1,  3, 5.                                                 | Pre-dose on  day 1, postdose on day  5, day 8 &  15                               |                                 |                                                    |                                            |                                |
| Day 8, 15d                                                                  |                               |                                 |                                                    |                                            |                                |
| 2                                                                           | 1                             | Pre-dose,                       |                                                    |                                            |                                |
| EOI, 1, 2, 3,  4, and 6  hours after  start of  LMB-100  infusion if  given | Pre-dose                      | Xb                              | Pre                                                    |                                            |                                |
| dose/EOI  on days  1, 3, 5                                                  | Pre-dose on  day 1, postdose on day  5, day 8 &  15                               |                                 |                                                    |                                            |                                |
| Day 8, 15 d                                                                 |                               |                                 |                                                    |                                            |                                |
| 3                                                                           | 1                             | X                               | Xb                                                 | Pre                                            |                                |
| dose/EOI                                                                    | Pre-dose  pembro              |                                 |                                                    |                                            |                                |
| 4                                                                           | 1                             | X                               | Xb                                                 | Pre                                            |                                |
| dose/EOI                                                                    | Pre-dose  pembro              |                                 |                                                    |                                            |                                |
| 5 and                                                                       | 1                             | Xb                              | Pre                                                    |                                            |                                |
| dose/EOI                                                                    |                               |                                 |                                                    |                                            |                                |
| beyond                                                                      | Pre-dose  pembro              |                                 |                                                    |                                            |                                |  
  
a. A fresh biopsy or tumor effusion sample will be collected if archival tumor tissue is not available.  
b. Optional tumor biopsies for research will be performed at baseline (C1D1), at the end of cycle 2 of LMB-100, after second cycle of pembrolizumab. In the event that patients get one cycle of LMB-100, optional tumor biopsies for research will be performed at baseline, at the end of cycle 1 of LMB-100 and after second cycle of pembrolizumab.  
c. Refers to the end of the first course. No additional research samples will be collected during the second course.  
d. If Day 15 assessments are performed at the NIH. Blood collection may occur -3/+1 day of C1D15 and C2D15.  
e. Samples may be collected 1 ± 3 days from indicated time.  
Note: Timed samples above must be drawn within a +/- 5 minute window for draws up to 4 hours post dose and within a +/- 20 minute window for 8-10 hr and 12-16 hr post dose draws.
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5.3 sample storage tracking and

Samples will be ordered in CRIS and tracked through a Clinical Trial Data Management system. Should a CRIS screen not be available, the CRIS downtime procedures will be followed. Samples will not be sent outside NIH without appropriate approvals and/or agreements, if required. 5.3.1 Blood Processing Core (BPC) Upon arrival in the Blood Processing Core (BPC), all samples are barcoded, with data entered and stored in Labmatrix, the system utilized by the CPP. This is a secure program, with access limited to defined Figg lab personnel, who are issued individual user accounts. Installation of Labmatrix is limited to computers specified by Dr. Figg. These computers have a password restricted login screen. Labmatrix creates a unique barcode ID for every sample and sample box, which cannot be traced back to patients without Labmatrix access. The data recorded for each sample includes the patient ID, name, trial name/protocol number, time drawn, cycle, time point, dose, material type, as well as box and freezer location. Patient demographics associated with the clinical center patient number are provided in the system. For each sample, there are notes associated with processing method (delay in sample processing, storage conditions on the ward, etc.).
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sample storage and destruction

Barcoded samples are stored in barcoded boxes in a locked freezer at either -20°C or -80°C according to stability requirements. These freezers are located onsite in the CPP and offsite at NCI Frederick Central Repository Services in Frederick, MD. Visitors to the laboratory are required to be accompanied by laboratory staff at all times. Access to stored clinical samples is restricted. Samples will be stored until requested by a researcher named on the protocol. All requests are monitored and tracked in Labmatrix. All researchers are required to sign a form stating that the samples are only to be used for research purposes associated with this trial (as per IRB approved protocol) and that any unused samples must be returned to the BPC. It is the responsibility of the NCI Principal Investigator to ensure that the samples requested are being used in a manner consistent with IRB approval. Following completion of this study, samples will remain in storage as detailed above. Access to these samples will only be granted following IRB approval of an additional protocol, granting the rights to use the material. If, at any time, a patient withdraws from the study and does not wish for their existing samples to be utilized, the individual must provide a written request. Following receipt of this request, the samples will be destroyed or returned to the patient, if so requested. The PI will record any loss or unanticipated destruction of samples as a deviation. Reporting will be per the requirements of section 7.2. Sample barcodes are linked to patient demographics and limited clinical information. This information will only be provided to investigators listed on this protocol, via registered use of the Labmatrix. It is critical that the sample remains linked to patient information such as race, age, dates of diagnosis and death, and histological information about the tumor, in order to correlate genotype with these variables.
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5.3.2 leidos biomedical research inc.

Blood and tissue collected during the course of this study will follow storage, handling and labeling procedures to ensure that security, confidentiality and sample integrity are maintained. All samples (blood or tissue) are tracked by distinct identification labels that include a unique patient identifier and date of specimen collection. Thus, samples will be coded, with access to the code key linking personal data restricted to the study investigators. All cryopreserved samples are tracked for freezer location and storage criteria. All Samples are stored in a locked freezer at -70C according to stability requirements. These freezers are located offsite at NCI-Frederick, at the Leidos Biomedical Research, Inc. Lab in Frederick, MD. Samples will be stored until requested by a researcher named on the protocol. All use and requests for use will be recorded by the Leidos Biomedical, Inc. Lab. Any unused samples must be returned. Some samples as indicated below may be stored in monitored freezers/refrigerators in the investigator's laboratory at specified temperatures with alarm systems in place. At the completion of this protocol, samples will remain in storage as detailed above. If additional studies are to be performed on any samples retaining patient identifiers, obtained during the conduct of this trial, access to these samples will only be granted following IRB approval of an additional protocol, granting the rights to use the material. If specimens are to be discarded at any point, they will be disposed of in accordance with the environmental protection laws, regulations and guidelines of the Federal Government and the State of Maryland. If, at any time, a patient withdraws from the study and does not wish for their existing samples to be utilized, the individual must provide a written request. Following receipt of this request, the samples will be destroyed (or returned to the patient, if so requested).
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5.3.3 nci laboratory of pathology

Tissues designated for clinical diagnostics are transported to the Laboratory of Pathology (LP) where they are examined grossly, and relevant portions are fixed, embedded in paraffin and sectioned and stained for diagnostic interpretation. Unutilized excess tissue that is not placed in paraffin blocks is stored in formalin for up to three months, in accordance with College of American Pathologists/Joint Commission on Accreditation of Healthcare Organizations (CAP/JCAHO) guidelines, and then discarded. Following completion of the diagnostic workup, the slides and tissue blocks are stored indefinitely in the LP's clinical archives. All specimens are catalogued and retrieved utilizing the clinical laboratory information systems, in accordance with CAP/JCAHO regulations. The use of any stored specimens for research purposes is only allowed when the appropriate IRB approval has been obtained. In some cases, this approval has been obtained via the original protocol on which the patient was enrolled.
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5.3.4 laboratory of dr. raffit

This study will follow storage, handling and labeling procedures to ensure that security, confidentiality and sample integrity are maintained. All samples (blood or tissue) are tracked by distinct identification labels generated by LabMatrix that include a unique patient identifier and date of specimen collection. Thus, samples will be coded, with access to the code key linking restricted to the study investigators. Depending on specimen type, samples are stored in liquid nitrogen, in monitored freezers/refrigerators at either -20 or -80C according to stability requirements or in a slide cabinet in the research Laboratory of Dr. Raffit Hassan (Building 10, Room 3B51)
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5.3.5 protocol completion/sample destruction

All specimens obtained in the protocol are used as defined in the protocol. Any specimens remaining at the completion of the protocol will be stored in the conditions described above. The study will remain open as long as sample or data analysis continues. Samples from consenting subjects will be stored until they are no longer of scientific value or until a subject withdraws consent for their continued use, at which time they will be destroyed. Once primary research objectives for the protocol are achieved, intramural researchers can request access to remaining samples, provided they have an IRB-approved protocol and patient consent or an exemption from IRB review. The PI will report any unanticipated loss or destruction of samples per section **7.2.1**. Freezer problems, lost samples, or other problems associated with samples that meet expedited reporting requirements (see section **7.2.1**) will also be reported.
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5.4 samples for genetic/genomic analysis

Using the tissue samples, we will perform the TruSight Oncology 500 Assay. This is a clinical next-generation sequencing-based multiple-biomarker assay developed by Thermo Fisher and targets most relevant single nucleotide variants, small insertions and deletions, copy number variants, and gene fusions from 161 cancer genes. 5.4.2 Privacy and Confidentiality of medical information/biological specimens Initially the samples of each patient will be barcoded. At no time will patient's names be used on the tissue samples. Sometimes, because a group collaboration or journal policy requires it, a subject's genetic data will be deposited in a database such as dbGaP. Although there is controlled access to such a database, such a submission carries theoretical risks of revealing the identity of the subject. This is discussed in the consent.
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5.4.3 management of results

Subjects will be contacted if a clinically actionable gene variant is discovered. Clinically actionable findings for the purpose of this study are defined as disorders appearing in the American College of Medical Genetics and Genomics recommendations for the return of incidental findings that is current at the time of primary analysis. (A list of current guidelines is maintained on the CCR intranet: https://ccrod.cancer.gov/confluence/display/CCRCRO/Incidental+Findings+Lists). Subjects will be contacted at this time with a request to provide a blood sample to be sent to a CLIA certified laboratory. The costs of CLIA testing will be paid for by the Center for Cancer Research, the Branch, or the Principal Investigator. If the health history, family history, or tumor diagnosis from the Laboratory of Pathology at the NIH Clinical Center suggests that the participant might benefit from genetic testing, we will discuss this with him/her.
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5.4.4 genetic counseling

If the research findings are verified in the CLIA certified lab, the subject will be offered the opportunity to come to NIH (at our expense) to have genetic education and counseling with the NCI Genetics Branch to explain this result. If the subject does not want to come to NIH, a referral to a local genetic healthcare provider will be provided (at their expense). This is the only time during the course of the study that incidental findings will be returned. No interrogations regarding clinically actionable findings will be made after the primary analysis.
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6 data collection and evaluation

The PI will be responsible for overseeing entry of data into an in-house password protected electronic system (C3D) and ensuring data accuracy, consistency and timeliness. The principal investigator, associate investigators/research nurses and/or a contracted data manager will assist with the data management efforts. Primary and final analyzed data will have identifiers so that research data can be attributed to an individual human subject participant. All adverse events, including clinically significant abnormal findings on laboratory evaluations, regardless of severity, will be followed until return to baseline or stabilization of event. Document AEs from the first study intervention, Study Day 1, through 90 days after the subject received the last product administration (first course and if applicable, second course). After 90 days (first course, and if applicable second course) only adverse events which are serious and related to the study investigational agent/intervention need to be recorded. An abnormal laboratory value will be recorded in the database as an AE **only** if the laboratory abnormality is characterized by any of the following: Results in discontinuation from the study  
Is associated with clinical signs or symptoms Requires treatment or any other therapeutic intervention Is associated with death or another serious adverse event, including hospitalization. Is judged by the Investigator to be of significant clinical impact If any abnormal laboratory result is considered clinically significant, the investigator will provide details about the action taken with respect to the test drug and about the patient's outcome.
End of study procedures: Data will be stored according to HHS, FDA regulations, and NIH Intramural Records Retention Schedule as applicable. Loss or destruction of data: Should we become aware that a major breach in our plan to protect subject confidentiality and trial data has occurred, this will be reported expeditiously per requirements in section **7.2.1**.  
6.2 DATA SHARING P**LANS**
6.2.1 Human Data Sharing Plan
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what data will be shared?

I will share human data generated in this research for future research as follows:  
Coded, linked data in an NIH-funded or approved public repository.  
Coded, linked data in BTRIS (automatic for activities in the Clinical Center)
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how and where will the

Data will be shared through: An NIH-funded or approved public repository: clinicaltrials.gov, dbGaP  
BTRIS (automatic for activities in the Clinical Center)
Publication and/or public presentations.
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when will the data be

Before publication. At the time of publication or shortly thereafter.  
6.2.2 Genomic Data Sharing Plan
Unlinked genomic data will be deposited in public genomic databases such as dbGaP in compliance with the NIH Genomic Data Sharing Policy.
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6.3 response criteria

For the purposes of this study, patients should be re-evaluated for response every 6 weeks (2 cycles). In addition to a baseline scan, confirmatory scans should also be obtained no less than 4 weeks following initial documentation of objective response. Refer to **Appendix C** for confirmatory scan and continued treatment instructions. Response and progression will be assessed by the investigator on the basis of physical examinations, computed tomography (CT) or Magnetic Resonance (MR) scans, and potentially other modalities according to standard of care. When the Investigator identifies radiographic progression while on pembrolizumab per RECIST 1.1 or mRECIST, efforts should be made to verify radiologic PD. Treatment should continue until PD has been verified. Regardless of whether PD is verified, if the Investigator considers the participant has progressed, but elects to implement iRECIST, the Investigator will assess for confirmation of progression by iRECIST at subsequent time points. When clinically stable, participants should not be discontinued until progression is confirmed by the Investigator, working with local radiology, according to the rules below. This allowance to continue treatment despite initial radiologic PD takes into account the observation that some participants can have a transient tumor flare in the first few months after the start of immunotherapy, and then experience subsequent disease response.
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6.3.1 disease parameters

Measurable disease: Measurable lesions are defined as those that can be accurately measured in at least one dimension (longest diameter to be recorded) as:
By chest x-ray: >20 mm;  
By CT scan:
o Scan slice thickness 5 mm or under: >10 mm o Scan slice thickness >5 mm: double the slice thickness With calipers on clinical exam: >10 mm.
All tumor measurements must be recorded in millimeters (or decimal fractions of centimeters). Malignant lymph nodes. To be considered pathologically enlarged and measurable, a lymph node must be >15 mm in short axis when assessed by CT scan (CT scan slice thickness recommended to be no greater than 5 mm). At baseline and in follow-up, only the short axis will be measured and followed. Non-measurable disease. All other lesions (or sites of disease), including small lesions (longest diameter <10 mm or pathological lymph nodes with ≥10 to <15 mm short axis), are considered non-measurable disease. Bone lesions, leptomeningeal disease, ascites, pleural/pericardial effusions, lymphangitis cutis/pulmonitis, inflammatory breast disease, and abdominal masses (not followed by CT or MRI), are considered as non-measurable. Note: Cystic lesions that meet the criteria for radiographically defined simple cysts should not be considered as malignant lesions (neither measurable nor non-measurable) since they are, by definition, simple cysts. 'Cystic lesions' thought to represent cystic metastases can be considered as measurable lesions, if they meet the definition of measurability described above. However, if non-cystic lesions are present in the same patient, these are preferred for selection as target lesions. Target lesions. All measurable lesions up to a maximum of 2 lesions per organ and 5 lesions in total, representative of all involved organs, should be identified as **target lesions** and recorded and measured at baseline. Target lesions should be selected on the basis of their size (lesions with the longest diameter), be representative of all involved organs, but in addition should be those that lend themselves to reproducible repeated measurements. It may be the case that, on occasion, the largest lesion does not lend itself to reproducible measurement in which circumstance the next largest lesion which can be measured reproducibly should be selected. A sum of the diameters (longest for non-nodal lesions, short axis for nodal lesions) for all target lesions will be calculated and reported as the baseline sum diameters. If lymph nodes are to be included in the sum, then only the short axis is added into the sum. The baseline sum diameters will be used as reference to further characterize any objective tumor regression in the measurable dimension of the disease. Non-target lesions. All other lesions (or sites of disease) including any measurable lesions over and above the 5 target lesions should be identified as **non-target lesions** and should also be recorded at baseline. Measurements of these lesions are not required, but the presence, absence, or in rare cases unequivocal progression of each should be noted throughout follow-up. 6.3.1.1 Methods for Evaluation of Measurable Disease All measurements should be taken and recorded in metric notation using a ruler or calipers. All baseline evaluations should be performed as closely as possible to the beginning of treatment and never more than 4 weeks before the beginning of the treatment. The same method of assessment and the same technique should be used to characterize each identified and reported lesion at baseline and during follow-up. Imaging-based evaluation is preferred to evaluation by clinical examination unless the lesion(s) being followed cannot be imaged but are assessable by clinical exam. Clinical lesions: Clinical lesions will only be considered measurable when they are superficial
(e.g., skin nodules and palpable lymph nodes) and 10 mm diameter as assessed using calipers
(e.g., skin nodules). In the case of skin lesions, documentation by color photography, including a ruler to estimate the size of the lesion, is recommended. Chest x-ray: Lesions on chest x-ray are acceptable as measurable lesions when they are clearly defined and surrounded by aerated lung. However, CT is preferable. Conventional CT and MRI: This guideline has defined measurability of lesions on CT scan based on the assumption that CT slice thickness is 5 mm or less. If CT scans have slice thickness greater than 5 mm, the minimum size for a measurable lesion should be twice the slice thickness. MRI is also acceptable in certain situations (e.g. for body scans). Use of MRI remains a complex issue. MRI has excellent contrast, spatial, and temporal resolution; however, there are many image acquisition variables involved in MRI, which greatly impact image quality, lesion conspicuity, and measurement. Furthermore, the availability of MRI is variable globally. As with CT, if an MRI is performed, the technical specifications of the scanning sequences used should be optimized for the evaluation of the type and site of disease. Furthermore, as with CT, the modality used at follow-up should be the same as was used at baseline and the lesions should be measured/assessed on the same pulse sequence. It is beyond the scope of the RECIST guidelines to prescribe specific MRI pulse sequence parameters for all scanners, body parts, and diseases. Ideally, the same type of scanner should be used and the image acquisition protocol should be followed as closely as possible to prior scans. Body scans should be performed with breath-hold scanning techniques, if possible. PET-CT: At present, the low dose or attenuation correction CT portion of a combined PET-CT is not always of optimal diagnostic CT quality for use with RECIST measurements. However, if the site can document that the CT performed as part of a PET-CT is of identical diagnostic quality to a diagnostic CT (with IV and oral contrast), then the CT portion of the PET-CT can be used for RECIST measurements and can be used interchangeably with conventional CT in accurately measuring cancer lesions over time. Note, however, that the PET portion of the CT introduces additional data, which may bias an investigator if it is not routinely or serially performed. Ultrasound: Ultrasound is not useful in assessment of lesion size and should not be used as a method of measurement. Ultrasound examinations cannot be reproduced in their entirety for independent review at a later date and, because they are operator dependent, it cannot be guaranteed that the same technique and measurements will be taken from one assessment to the next. If new lesions are identified by ultrasound in the course of the study, confirmation by CT or MRI is advised. If there is concern about radiation exposure at CT, MRI may be used instead of CT in selected instances. Endoscopy, Laparoscopy: The utilization of these techniques for objective tumor evaluation is not advised. However, such techniques may be useful to confirm complete pathological response when biopsies are obtained or to determine relapse in trials where recurrence following complete response (CR) or surgical resection is an endpoint.  
Tumor markers: Tumor markers alone cannot be used to assess response. If markers are initially above the upper normal limit, they must normalize for a patient to be considered in complete clinical response. Specific guidelines for both CA-125 response (in recurrent ovarian cancer) and PSA response (in recurrent prostate cancer) have been published.(42-44) In addition, the Gynecologic Cancer Intergroup has developed CA-125 progression criteria which are to be integrated with objective tumor assessment for use in first-line trials in ovarian cancer.(45) Cytology, Histology: These techniques can be used to differentiate between partial responses (PR) and complete responses (CR) in rare cases (e.g., residual lesions in tumor types, such as germ cell tumors, where known residual benign tumors can remain). The cytological confirmation of the neoplastic origin of any effusion that appears or worsens during treatment when the measurable tumor has met criteria for response or stable disease is mandatory to differentiate between response or stable disease (an effusion may be a side effect of the treatment) and progressive disease. FDG-PET: While FDG-PET response assessments need additional study, it is sometimes reasonable to incorporate the use of FDG-PET scanning to complement CT scanning in assessment of progression (particularly possible 'new' disease). New lesions on the basis of FDG-PET imaging can be identified according to the following algorithm: a. Negative FDG-PET at baseline, with a positive FDG-PET at follow-up is a sign of PD based on a new lesion. b. No FDG-PET at baseline and a positive FDG-PET at follow-up: If the positive FDG-PET
at follow-up corresponds to a new site of disease confirmed by CT, this is PD. If the positive FDGPET at follow-up is not confirmed as a new site of disease on CT, additional follow-up CT scans are needed to determine if there is truly progression occurring at that site (if so, the date of PD will be the date of the initial abnormal FDG-PET scan). If the positive FDG-PET at follow-up corresponds to a pre-existing site of disease on CT that is not progressing on the basis of the anatomic images, this is not PD. c. FDG-PET may be used to upgrade a response to a CR in a manner similar to a biopsy in cases where a residual radiographic abnormality is thought to represent fibrosis or scarring. The use of FDG-PET in this circumstance should be prospectively described in the protocol and supported by disease-specific medical literature for the indication. However, it must be acknowledged that both approaches may lead to false positive CR due to limitations of FDG-PET and biopsy resolution/sensitivity. Note: A 'positive' FDG-PET scan lesion means one which is FDG avid with an uptake greater than twice that of the surrounding tissue on the attenuation corrected image. 6.3.1.2 RECIST version 1.1 Response Criteria 6.3.1.2.1 Evaluation of Target Lesions Complete Response (CR): Disappearance of all target lesions. Any pathological lymph nodes (whether target or non-target) must have reduction in short axis to <10 mm. Partial Response (PR): At least a 30% decrease in the sum of the diameters of target lesions, taking as reference the baseline sum of diameters. Progressive Disease (PD): At least a 20% increase in the sum of the diameters of target lesions, taking as reference the smallest sum on study (this includes the baseline sum if that is the smallest on study). In addition to the relative increase of 20%, the sum must also demonstrate an absolute increase of at least 5 mm. (Note: the appearance of one or more new lesions is also considered progressions). Stable Disease (SD): Neither sufficient shrinkage to qualify for PR nor sufficient increase to qualify for PD, taking as reference the smallest sum of diameters while on study.  
6.3.1.2.2 Evaluation of Non-Target Lesions Complete Response (CR): Disappearance of all non-target lesions and normalization of tumor marker level. All lymph nodes must be non-pathological in size (<10 mm short axis). Note: If tumor markers are initially above the upper normal limit, they must normalize for a patient to be considered in complete clinical response. Non-CR/Non-PD: Persistence of one or more non-target lesion(s) and/or maintenance of tumor marker level above the normal limits. Progressive Disease (PD): Appearance of one or more new lesions and/or *unequivocal progression* of existing non-target lesions. *Unequivocal progression* should not normally trump target lesion status. It must be representative of overall disease status change, not a single lesion increase. Although a clear progression of "non-target" lesions only is exceptional, the opinion of the treating physician should prevail in such circumstances, and the progression status should be confirmed at a later time by the review panel (or Principal Investigator).  
6.3.1.2.3 Evaluation of Best Overall Response The best overall response is the best response recorded from the start of the treatment until disease progression/recurrence (taking as reference for progressive disease the smallest measurements recorded since the treatment started). The patient's best response assignment will depend on the achievement of both measurement and confirmation criteria.  
| For Patients with Measurable Disease (i.e., Target Disease) Target  Non-Target  New  Overall  Best Overall Response when  Lesions Lesions Lesions Response Confirmation is Required* CR CR No CR >4 wks. Confirmation** CR NonCR/Non-PD No PR CR Not  No PR evaluated >4 wks. Confirmation** PR NonCR/NonPD/not  evaluated No PR SD NonCR/NonPD/not  evaluated No SD Documented at least once >4  wks. from baseline** PD Any Yes or  PD No Any PD*** Yes or  PD No no prior SD, PR or CR Any Any Yes PD       |                                                                                                                      |
|-------|----------------------------------------------------------------------------------------------------------------------|
| *     | See RECIST 1.1 manuscript for further details on what is evidence of a new lesion.                                   |
| **    | Only for non-randomized trials with response as primary endpoint.                                                    |
| ***   | In exceptional circumstances, unequivocal progression in non-target lesions may be accepted as disease  progression. |
| Note: | Patients with a global deterioration of health status requiring discontinuation of treatment without objective       |  
*** In exceptional circumstances, unequivocal progression in non-target lesions may be accepted as disease progression.  
Note: Patients with a global deterioration of health status requiring discontinuation of treatment without objective evidence of disease progression at that time should be reported as "*symptomatic deterioration."* Every effort should be made to document the objective progression even after discontinuation of treatment.  
For Patients with Non-Measurable Disease (i.e., Non-Target Disease)  
| Non-Target Lesions   | New Lesions   | Overall Response   |
|----------------------|---------------|--------------------|
| CR                   | No            | CR                 |
| Non-CR/non-PD        | No            | Non-CR/non-PD*     |  
| Non-Target Lesions                                                                                                                                                                                                                             | New Lesions   | Overall Response   |
|------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|---------------|--------------------|
| Not all evaluated                                                                                                                                                                                                                              | No            | not evaluated      |
| Unequivocal PD                                                                                                                                                                                                                                 | Yes or No     | PD                 |
| Any                                                                                                                                                                                                                                            | Yes           | PD                 |
| * 'Non-CR/non-PD' is preferred over 'stable disease' for non-target disease since SD is increasingly used as  an endpoint for assessment of efficacy in some trials so to assign this category when no lesions can be  measured is not advised |               |                    |
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6.3.2 irecist

6.3.2.1 Assessment at Screening and Prior to RECIST 1.1 Progression Until radiographic progression based on RECIST 1.1, there is no distinct iRECIST assessment. 6.3.2.2 Assessment and Decision at RECIST 1.1 Progression In participants who show evidence of radiological PD by RECIST 1.1 the Investigator will decide whether to continue a participant on study treatment until repeat imaging is obtained (using iRECIST for participant management (see **Appendix C**). This decision by the Investigator should be based on the participant's overall clinical condition. Clinical stability is defined as the following:
Absence of symptoms and signs indicating clinically significant progression of disease  
No decline in ECOG performance status No requirements for intensified management, including increased analgesia, radiation, or other palliative care
Any participant deemed clinically unstable should be discontinued from study treatment at siteassessed first radiologic evidence of PD and is not required to have repeat tumor imaging for confirmation of PD by iRECIST.  
If the Investigator decides to continue treatment, the participant may continue to receive study treatment and the tumor assessment should be repeated 4 to 8 weeks later to confirm PD by iRECIST, per Investigator assessment. Tumor flare may manifest as any factor causing radiographic progression per RECIST 1.1, including:
Increase in the sum of diameters of target lesion(s) identified at baseline to ≥ 20% and
≥ 5 mm from nadir o Please note: the iRECIST publication uses the terminology "sum of measurements", but
"sum of diameters" will be used in this protocol, consistent with the original RECIST 1.1 terminology.  
Unequivocal progression of non-target lesion(s) identified at baseline  
Development of new lesion(s)
iRECIST defines new response categories, including iUPD (unconfirmed progressive disease) and iCPD (confirmed progressive disease). For purposes of iRECIST assessment, the first visit showing progression according to RECIST 1.1 will be assigned a visit (overall) response of iUPD, regardless of which factors caused the progression. At this visit, target and non-target lesions identified at baseline by RECIST 1.1 will be assessed as usual. New lesions will be classified as measurable or non-measurable, using the same size thresholds and rules as for baseline lesion assessment in RECIST 1.1. From measurable new lesions, up to 5 lesions total (up to 2 per organ), may be selected as New Lesions - Target. The sum of diameters of these lesions will be calculated and kept distinct from the sum of diameters for target lesions at baseline. All other new lesions will be followed qualitatively as New Lesions - Non-target. 6.3.2.3 Assessment at the Confirmatory Imaging On the confirmatory imaging, the participant will be classified as progression confirmed (with an overall response of iCPD), or as showing persistent unconfirmed progression (with an overall response of iUPD), or as showing disease stability or response (iSD/iPR/iCR). 6.3.2.4 Confirmation of Progression Progression is considered confirmed, and the overall response will be iCPD, if ANY of the following occurs:
Any of the factors that were the basis for the initial iUPD show worsening o For target lesions, worsening is a further increase in the sum of diameters of ≥ 5 mm, compared to any prior iUPD time point o For non-target lesions, worsening is any significant growth in lesions overall, compared to a prior iUPD time point; this does not have to meet the "unequivocal" standard of RECIST 1.1  
o For new lesions, worsening is any of these:
An increase in the new lesion sum of diameters by ≥ 5 mm from a prior iUPD time point Visible growth of new non-target lesions The appearance of additional new lesions Any new factor appears that would have triggered PD by RECIST 1.1 6.3.2.5 Persistent iUPD
Progression is considered not confirmed, and the overall response remains iUPD, if:
None of the progression-confirming factors identified above occurs AND  
The target lesion sum of diameters (initial target lesions) remains above the initial PD
threshold (by RECIST 1.1)
Additional imaging for confirmation should be scheduled 4 to 8 weeks from the scan on which iUPD is seen. This may correspond to the next visit in the original visit schedule. The assessment of the subsequent confirmation scan proceeds in an identical manner, with possible outcomes of iCPD, iUPD, and iSD/iPR/iCR.
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6.3.2.6 resolution of iupd

Progression is considered not confirmed, and the overall response becomes iSD/iPR/iCR, if:  
None of the progression-confirming factors identified above occurs, AND
The target lesion sum of diameters (initial target lesions) is not above the initial PD
threshold.
The response is classified as iSD or iPR (depending on the sum of diameters of the target lesions), or iCR if all lesions resolve. In this case, the initial iUPD is considered to be pseudo-progression, and the level of suspicion for progression is "reset". This means that the next visit that shows radiographic progression, whenever it occurs, is again classified as iUPD by iRECIST, and the confirmation process is repeated before a response of iCPD can be assigned.  
6.3.2.6.1 Management Following the Confirmatory Imaging If repeat imaging does not confirm PD per iRECIST, as assessed by the Investigator, and the participant continues to be clinically stable, study treatment may continue and follow the regular imaging schedule. If PD is confirmed, participants will be discontinued from study treatment. NOTE: If a participant has confirmed radiographic progression (iCPD) as defined above, but the participant is achieving a clinically meaningful benefit, an exception to continue study treatment may be considered. In this case, if study treatment is continued, tumor imaging should continue to be performed following the intervals as outlined in section 6.  
6.3.2.6.2 Detection of Progression at Visits After Pseudo-Progression Resolves After resolution of pseudo-progression (i.e., achievement of iSD/iPR/iCR), iUPD is indicated by any of the following events:
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target lesions

o Sum of diameters reaches the PD threshold (≥ 20% and ≥ 5 mm increase from nadir)
either for the first time, or after resolution of previous pseudo-progression. The nadir is always the smallest sum of diameters seen during the entire trial, either before or after an instance of pseudo-progression.  
Non-target lesions o If non-target lesions have never shown unequivocal progression, their doing so for the first-time results in iUPD.  
o If non-target lesions had shown previous unequivocal progression, and this progression has not resolved, iUPD results from any significant further growth of non-target lesions, taken as a whole.  
New lesions o New lesions appear for the first time  
o Additional new lesions appear
o Previously identified new target lesions show an increase of ≥ 5 mm in the new lesion sum of diameters, from the nadir value of that sum o Previously identified non-target lesions show any significant growth If any of the events above occur, the overall response for that visit is iUPD, and the iUPD evaluation process (see Assessment at the Confirmatory Imaging above) is repeated. Progression must be confirmed before iCPD can occur. The decision process is identical to the iUPD confirmation process for the initial PD, except in one respect. If new lesions occurred at a prior instance of iUPD, and at the confirmatory scan the burden of new lesions has increased from its smallest value (for new target lesions, their sum of diameters is ≥5 mm increased from its nadir), then iUPD cannot resolve to iSD or iPR. It will remain iUPD until either a decrease in the new lesion burden allows resolution to iSD or iPR, or until a confirmatory factor causes iCPD. Additional details about iRECIST are provided in the iRECIST publication.(46) 6.3.3 Duration of Response Duration of overall response: The duration of overall response is measured from the time measurement criteria are met for CR or PR (whichever is first recorded) until the first date that recurrent or progressive disease is objectively documented (taking as reference for progressive disease the smallest measurements recorded since the treatment started). The duration of overall CR is measured from the time measurement criteria are first met for CR until the first date that progressive disease is objectively documented. Duration of stable disease: Stable disease is measured from the start of the treatment until the criteria for progression are met, taking as reference the smallest measurements recorded since the treatment started, including the baseline measurements. 6.3.4 Progression-Free Survival Progression free survival (PFS) is defined as the duration of time from start of treatment to time of progression (on or after pembrolizumab) or death, whichever occurs first. 6.3.5 Objective Response Rate Objective response rate (ORR) is defined as the proportion of patients with partial response or complete response.
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6.4 toxicity criteria

The following adverse event management guidelines are intended to ensure the safety of each patient while on the study. The descriptions and grading scales found in the revised NCI Common Terminology Criteria for Adverse Events (CTCAE) version 5.0 will be utilized for AE reporting. All appropriate treatment areas should have access to a copy of the CTCAE version 5.0. A copy of the CTCAE version 5.0 can be downloaded from the CTEP web site (http://ctep.cancer.gov/protocolDevelopment/electronic_applications/ctc.htm)
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7 nih reporting requirements/data safety

Please refer to definitions provided in Policy 801: Reporting Research Events found here.
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7.2 ohsrp office of compliance

Please refer to the reporting requirements in **Policy 801: Reporting Research Events** and Policy 802 Non-Compliance Human Subjects Research found here. Note: Only IND Safety Reports that meet the definition of an unanticipated problem will need to be reported per these policies. 7.2.2 IRB Requirements for PI Reporting at Continuing Review Please refer to the reporting requirements in Policy 801: Reporting Research Events found here.
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7.2.3 nci clinical director reporting

Problems expeditiously reported to the OHSRP in iRIS will also be reported to the NCI Clinical Director. A separate submission is not necessary as reports in iRIS will be available to the Clinical Director. In addition to those reports, deaths not reported to the OHSRP that occur within 30 days after receiving a research intervention should be reported via email to the Clinical Director unless they are due to progressive disease. To report these deaths, please send an email describing the circumstances of the death to Dr. Dahut at NCICCRQA@mail.nih.gov within one business day of learning of the death.
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7.3 nih required data and

The clinical research team will meet on a regular basis weekly when patients are being actively treated on the trial to discuss each patient. All data will be collected in a timely manner and reviewed by the principal investigator or a lead associate investigator. Events meeting requirements for expedited reporting as described in section 7.2.1 will be submitted within the appropriate timelines. The principal investigator will review adverse event and response data on each patient to ensure safety and data accuracy. The principal investigator will personally conduct or supervise the investigation and provide appropriate delegation of responsibilities to other members of the research staff. 7.3.2 Safety Monitoring Committee (SMC) This protocol will be periodically reviewed by an intramural Safety Monitoring Committee. Initial review will occur as soon as possible after the annual NIH Intramural IRB continuing review date. Subsequently, each protocol will be reviewed as close to annually as the quarterly meeting schedule permits or more frequently as may be required by the SMC based on the risks presented in the study. For initial and subsequent reviews, protocols will not be reviewed if there is no accrual within the review period. The SMC review will focus on unexpected protocol-specific safety issues that are identified during the conduct of the clinical trial. Written outcome letters will be generated in response to the monitoring activities and submitted to the Principal investigator and Clinical Director or Deputy Clinical Director, CCR, NCI.
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8 sponsor safety reporting 8.1

Any untoward medical occurrence in a patient or clinical investigation subject administered a pharmaceutical product and which does not necessarily have a causal relationship with this treatment. An adverse event (AE) can therefore be any unfavorable and unintended sign (including an abnormal laboratory finding), symptom, or disease temporally associated with the use of a medicinal (investigational) product, whether or not related to the medicinal (investigational) product (ICH E6 (R2)) 8.1.2 Serious Adverse Event (SAE) An adverse event or suspected adverse reaction is considered serious if in the view of the investigator or the sponsor, it results in any of the following:
Death,  
A life-threatening adverse event (see **8.1.3**) Inpatient hospitalization or prolongation of existing hospitalization o A hospitalization/admission that is pre-planned (i.e., elective or scheduled surgery arranged prior to the start of the study), a planned hospitalization for pre-existing condition, or a procedure required by the protocol, without a serious deterioration in health, is not considered a serious adverse event.  
o A hospitalization/admission that is solely driven by non-medical reasons (e.g.,
hospitalization for patient convenience) is not considered a serious adverse event.  
o Emergency room visits or stays in observation units that do not result in admission to the hospital would not be considered a serious adverse event. The reason for seeking medical care should be evaluated for meeting one of the other serious criteria.  
Persistent or significant incapacity or substantial disruption of the ability to conduct normal life functions A congenital anomaly/birth defect Important medical events that may not result in death, be life-threatening, or require hospitalization may be considered a serious adverse drug experience when, based upon appropriate medical judgment, they may jeopardize the patient or subject and may require medical or surgical intervention to prevent one of the outcomes listed in this definition.  
8.1.3 Life-threatening
An adverse event or suspected adverse reaction is considered "life-threatening" if, in the view of either the investigator or sponsor, its occurrence places the patient or subject at immediate risk of death. It does not include an adverse event or suspected adverse reaction that, had it occurred in a more severe form, might have caused death. (21CFR312.32)
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8.1.4 severity

The severity of each Adverse Event will be assessed utilizing the CTCAE version 5. 8.1.5 Relationship to Study Product All AEs will have their relationship to study product assessed using the terms: related or not related.  
Related - There is a reasonable possibility that the study product caused the adverse event. Reasonable possibility means that there is evidence to suggest a causal relationship between the study product and the adverse event.  
Not Related - There is not a reasonable possibility that the administration of the study product caused the event.
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8.2 assessment of safety events

AE information collected will include event description, date of onset, assessment of severity and relationship to study product and alternate etiology (if not related to study product), date of resolution of the event, seriousness and outcome. The assessment of severity and relationship to the study product will be done only by those with the training and authority to make a diagnosis and listed on the Form FDA 1572 as the site principal investigator or sub-investigator. AEs occurring during the collection and reporting period will be documented appropriately regardless of relationship. AEs will be followed through resolution.
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saes will be

Assessed for severity and relationship to study product and alternate etiology (if not related to study product) by a licensed study physician listed on the Form FDA 1572 as the site principal investigator or sub-investigator.  
Recorded on the appropriate SAE report form, the medical record and captured in the clinical database.  
Followed through resolution by a licensed study physician listed on the Form FDA 1572 as the site principal investigator or sub-investigator.
For timeframe of recording adverse events, please refer to section 6.1. All serious adverse events recorded from the time of first investigational product administration must be reported to the sponsor with the exception of any listed in section 8.4.
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8.3 reporting of serious adverse

Any AE that meets protocol-defined serious criteria or meets the definition of Adverse Event of Special Interest that require expedited reporting must be submitted immediately (within 24 hours of awareness) to OSRO Safety using the CCR SAE report form. Any exceptions to the expedited reporting requirements are found in section 8.4. All SAE reporting must include the elements described in 8.2. SAE reports will be submitted to the Center for Cancer Research (CCR) at: OSROSafety@mail.nih.gov and to the CCR PI and study coordinator. CCR SAE report form and instructions can be found at: https://ccrod.cancer.gov/confluence/display/CCRCRO/Forms+and+Instructions Following the assessment of the SAE by OSRO, other supporting documentation of the event may be requested by the OSRO Safety and should be provided as soon as possible.
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8.4 waiver of expedited reporting

As death due to disease progression is a part of the study objectives (PFS and OS) and captured as an endpoint in this study, they will not be reported in expedited manner to the Sponsor. However, if there is evidence suggesting a causal relationship between the study drug and the event, report the event in an expedited manner according to section 8.3. Hospitalization that is deemed to be due to disease progression, and not attributable to the intervention will not be reported as an SAE. The event, and the assessment that it was caused by disease progression will be documented in the medical records. The causality assessment of hospitalization will be re‐evaluated any time when new information is received. If the causality assessment changes from disease progression to related to the study intervention, SAE report will be sent to the Sponsor in an expedited manner according to Section 8.3. If there is any uncertainty whether the intervention is a contributing factor to the event, the event should be reported as AE or SAE as appropriate.
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8.5 reporting pregnancy

All required pregnancy reports/follow-up to OSRO will be submitted to: OSROSafety@mail.nih.gov and to the CCR PI and study coordinator. Forms and instructions can be found here: https://ccrod.cancer.gov/confluence/display/CCRCRO/Forms+and+Instructions
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8.5.1 maternal exposure

If a patient becomes pregnant during the course of the study, the study treatment should be discontinued immediately, and the pregnancy reported to the Sponsor no later than 24 hours of when the Investigator becomes aware of it. The Investigator should notify the Sponsor no later than 24 hours of when the outcome of the Pregnancy become known, Pregnancy itself is not regarded as an SAE. However, congenital abnormalities or birth defects and spontaneous miscarriages that meet serious criteria (**8.1.2**) should be reported as SAEs. The outcome of all pregnancies (spontaneous miscarriage, elective termination, ectopic pregnancy, normal birth, or congenital abnormality) should be followed up and documented. 8.5.2 Paternal exposure Male patients should refrain from fathering a child or donating sperm during the study and for 180 days after the last dose of LMB-100 or pembrolizumab. Pregnancy of the patient's partner is not considered to be an AE. However, the outcome of all pregnancies (spontaneous miscarriage, elective termination, ectopic pregnancy, normal birth, or congenital abnormality) occurring from the date of the first dose until 180 days after the last dose should, if possible, be followed up and documented.
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8.6 regulatory reporting for studies

Following notification from the investigator, CCR, the IND sponsor, will report any suspected adverse reaction that is both serious and unexpected. CCR will report an AE as a suspected adverse reaction only if there is evidence to suggest a causal relationship between the study product and the adverse event. CCR will notify FDA and all participating investigators (i.e., all investigators to whom the sponsor is providing drug under its INDs or under any investigator's IND) in an IND safety report of potential serious risks from clinical trials or any other source, as soon as possible, in accordance to 21 CFR Part 312.32. All serious events will be reported to the FDA at least annually in a summary format.
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9 clinical monitoring

As a sponsor for clinical trials, FDA regulations require the CCR to maintain a monitoring program. The CCR's program allows for confirmation of: study data, specifically data that could affect the interpretation of primary and secondary study endpoints; adherence to the protocol, regulations, ICH E6, and SOPs; and human subjects protection. This is done through independent verification of study data with source documentation focusing on:
Informed consent process Eligibility confirmation Drug administration and accountability Adverse events monitoring Response assessment The monitoring program also extends to multi-site research when the CCR is the coordinating center. This trial will be monitored by personnel employed by a CCR contractor. Monitors are qualified by training and experience to monitor the progress of clinical trials. Personnel monitoring this study will not be affiliated in any way with the trial conduct.
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10 statistical considerations 10.1 statistical

10.1.1 Primary Endpoint
To determine the objective response rate for the combination of LMB-100 and Pembrolizumab in the treatment of patients with advanced, mesothelin-expressing nonsquamous non-small cell lung cancer (NSCLC) previously treated with immune checkpoint inhibitors.  
10.1.2 Secondary Endpoints To determine the overall survival (OS), duration of response and progression free survival
(PFS) of LMB-100 and Pembrolizumab in the treatment of patients with advanced, nonsquamous non-small cell lung cancer (NSCLC).  
To determine the safety and tolerability of combination of LMB-100 and Pembrolizumab in the treatment of patients with advanced, mesothelin-expressing non-squamous nonsmall cell lung cancer (NSCLC) previously treated with immune checkpoint inhibitors 10.1.3 Sample size determination:
Patients will be screened for adequate mesothelin expression. Patients eligible for treatment will directly enrolled without randomization or any stratification. An observed response rate of 10% or greater would be considered desirable in this population of subjects who have previously failed a checkpoint inhibitor. In this arm, the trial will be conducted using a Simon minimax two-stage phase II trial design (47) to rule out an unacceptably low response rate of 5% (p0=0.05) in favor of an improved response rate of 20% (p1=0.20). With alpha=0.10 (probability of accepting a poor treatment=0.10) and beta = 0.20 (probability of rejecting a good treatment=0.20), the first stage will enroll 12 evaluable patients, and if 0 of the 12 have a response, then no further patients will be accrued. If 1 or more of the first 12 patients have a response, then accrual would continue until a total of 21 evaluable patients have been treated on Arm A. As it may take up to several months to determine if a patient has experienced a response, a temporary pause in the accrual may be necessary to ensure that enrollment to the second stage is warranted. If there are 1-2 patients with a response out of 21 patients, this would be an uninterestingly low response rate. If there were 3 or more of 21 (14.3%) who experienced a response, this would be sufficiently interesting to warrant further study in later trials. Under the null hypothesis (5% response rate), the probability of early termination is 54.0%. It is expected that 25% of patients who have NSCLC will have adequate (25%) mesothelin expression to receive treatment on this protocol and 10% of patient with lung adenocarcinoma coexpress PD-L1 and >25% mesothelin expression.  
In order to enroll and treat up to 21 evaluable patients with adequate mesothelin expression, the trial will plan to enroll and screen up to 100 patients. With 100 patients, there would be 85.1% probability of enrolling 21 or more evaluable patients if the true probability of adequate expression for a given patient was 25%. It is expected that up to 2 patients per month may be screened on this protocol, yielding approximately 5 to 6 treated patients per year. It is expected that up to 4 years may be required to enroll up to 100 patients for screening and 21 evaluable for response. To allow for a small number of unevaluable patients and screen failures 23 patients will be treated and the screening will be set at 100 total patients.
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10.2 populations for analyses 10.2.1

All patients will be evaluable for toxicity from the time of their first treatment with LMB-100. Events occurring after initiation of a second course of therapy will not be included in this analysis. 10.2.2 Evaluable for objective response Only those patients who have measurable disease present at baseline, have received at least one cycle of therapy, and have had their disease re-evaluated will be considered evaluable for response. Patients on the second course will not be included in this analysis
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10.3 statistical analyses 10.3.1 general

Efficacy will be assessed as the fraction of patients who experience a response and will be reported along with confidence intervals. Safety and tolerability will be assessed in a descriptive fashion. 10.3.2 Analysis of Primary Endpoint (s)
The fraction of patients who experience a response will be reported along with 80% and 95% twosided confidence intervals.
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10.3.3 analysis of the secondary

Overall Survival, and progression free survival will be calculated from the on-study date using the Kaplan-Meier method. Duration of response will be calculated for responders only, beginning at the date a patient is noted to have at least a PR. Safety and tolerability of LMB-100 + pembrolizumab will be assessed by reporting the grade of adverse events noted in each patient and reporting the proportion with grade 3 and grade 4 adverse events. The summary of the proportion with grade 3 and 4 adverse events will constitute the main analysis of the safety data. In addition, the number of patients who are treated who are unable to tolerate the treatment for other than development of adverse events will be reported.
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10.3.4 safety analyses

Safety and tolerability of LMB-100 + pembrolizumab, will be assessed by reporting the grade of adverse events noted in each patient and reporting the proportion with grade 3 and grade 4 adverse events. The summary of the proportion with grade 3 and 4 adverse events will constitute the main analysis of the safety data. Safety data will be presented in individual listings. Summaries will also be prepared. The safety data will consist of the reporting of all adverse events, vital signs, physical examination data, and appropriate laboratory safety data. 10.3.5 Baseline Descriptive Statistics Baseline demographic characteristics will be reported. 10.3.6 Planned Interim Analyses As indicated in the two-stage design, the number of responses after 12 evaluable have been treated will be noted and will be used to determine if enrollment to the second stage of accrual may proceed. 10.3.7 Sub-Group Analyses None 10.3.8 Tabulation of individual Participant Data None. 10.3.9 Exploratory Analyses The following are the exploratory analyses which may be undertaken:
To establish the correlation of response with tumor mesothelin expression.  
To evaluate correlation of tumor response with tumor PD-L1 expression.  
To evaluate changes in the tumor microenvironment following treatment with LMB100 followed by pembrolizumab.  
To evaluate the utility of serum mesothelin and megakaryocyte potentiating factor
(MPF) as a biomarkers of tumor response.  
To define the pharmacokinetics characteristics of LMB-100 to correlate responses with LMB-100 blood levels.  
To determine the incidence of antibody development at the end of cycles 1 and 2 with this combination.  
Correlation of incidence of KRAS mutation with mesothelin immunohistochemical expression in tumor samples.  
To determine the utility of circulating tumor DNA (ctDNA) and characteristic mutations for biomarkers analysis of response in blood, and overall tumor genome evolution over the course of treatment.  
To determine if there is association between PD-L1 expression level or mesothelin expression level and response, patients will be divided into responders vs. nonresponders and these expression levels will be compared between the two groups using an exact Wilcoxon rank sum test.  
To assess changes in circulating immune cell and cytokine levels caused by the treatment
Any of the exploratory evaluations which generate quantitative measures will be done using descriptive statistics including confidence intervals when appropriate. Any statistical tests performed for evaluation of exploratory objectives will be done without formal adjustment for multiple comparisons, but in the context of the number of tests performed.
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11 human subjects protections 11.1

LMB-100 is a mesothelin-targeted cytolytic fusion protein (cFP) and has shown preclinical dosedependent activity in monotherapy and/or combination in xenografts representing MSLN-positive indications (NSCLC, mesothelioma, triple negative breast cancer, gastric cancer, pancreas, ovarian, potentially other tumor indications). LMB-100 has shown synergy in laboratory models with inhibitors of the PD1 axis. Pembrolizumab is an anit-PD1/PD-L1 inhibitor. The rationale to evaluate LMB-100 with pembrolizumab in mesothelin expressing NSCLC patients is to determine the effect of the addition of pembrolizumab to LMB-100 therapy. All patients meeting the criteria listed in section **2.1** are eligible for enrollment.
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11.2 participation of children

There are no dosing or adverse event data are currently available on the use of LMB-100 with pembrolizumab in patients <18 years of age; therefore, children are excluded from this study.
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11.3 participation of subjects unable

Adults unable to give consent are excluded from enrolling in the protocol. However, re-consent may be necessary and there is a possibility, though unlikely, that subjects could become decisionally impaired. For this reason and because there is a prospect of direct benefit from research participation (section **11.5**), all subjects ≥ age 18 will be offered the opportunity to fill in their wishes for research and care, and assign a substitute decision maker on the "NIH Advance Directive for Health Care and Medical Research Participation" form so that another person can make decisions about their medical care in the event that they become incapacitated or cognitively impaired during the course of the study. Note: The PI or AI will contact the NIH Ability to Consent Assessment Team (ACAT) for evaluation as needed for the following: an independent assessment of whether an individual has the capacity to provide consent; assistance in identifying and assessing an appropriate surrogate when indicated; and/or an assessment of the capacity to appoint a surrogate. Please see section **11.6.1** for consent procedure.
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11.4 evaluation of benefits and

Patient safety will be managed by careful proactive patient selection prior to study to exclude patients at risk from study treatment due to their pre-existing conditions. During the study, safety of patients will be proactively managed by protocol-mandated physical examinations, vital signs assessments, ECGs, clinical laboratory assessments, and collection of adverse events and their assessment.  
The risks of the study include those associated with the study agents as discussed in section 13.  
11.4.2 Blood Collection Side effects of blood draws include pain and bruising, lightheadedness, and rarely, fainting. 11.4.3 Biopsy Collection The risks of the research biopsies collected at baseline, Post-C2, and after Pembrolizumab include pain, bleeding and infection at the biopsy site. 11.4.4 CT contrast In addition to the radiation risks from the scans discussed above, patients may experience an allergic reaction to CT contrast dye.
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11.4.5 radiation risks

The biopsies referenced above will be collected under CT guidance and subjects will have up to 9 CTs of the chest/abdomen/pelvis and 9 FDG PET/CT scans in a year. Subjects in this study may be exposed to approximately 23.9 rem total. 11.4.6 Non-Physical Risks of Genetic Research
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11.4.6.1 risk of receiving unwanted

Anxiety and stress may arise as a result of the anticipation that unwanted information regarding disease related DNA sequencing or disease tendencies, or misattributed paternity. Patients will be clearly informed that the data related to DNA sequencing and genetic analysis is coded, investigational and will not be shared with patients, family members or health care providers. 11.4.6.2 Risk related to possibility that information may be released This includes the risk that data related to genotype, DNA sequencing or risk for disease tendency or trait can be released to members of the public, insurers, employers, or law enforcement agencies. Although there are no plans to release results to the patients, family members or health care providers, this risk will be included in the informed consent document.
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11.4.6.3 risk to family or

Family members or relatives may or may not want to be aware of familial tendencies or genetic risks of disease which may cause anxiety about possible future health problems. As previously noted, patients will be notified of any medically significant and actionable incidental findings. Study results will not be shared with patients.
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11.5 risks/benefits analysis

Patients with advanced NSCLC are in continuous need of improved therapy options. This is especially true for patients where no standard therapy exists such as the patient population that will be eligible for this trial. Preclinical data has demonstrated promising anti-tumor efficacy of LMB100 in xenograft models in monotherapy and combination therapy. Laboratory studies have further demonstrated synergy with PD1/PD-L1 inhibitors. Therefore, LMB-100 + pembrolizumab may improve clinical outcome of patients with NSCLC. A number of clinically appropriate strategies to minimize risk to patients have been built into the protocol through the means of inclusion/exclusion criteria, monitoring strategies, and management guidelines. Overall, the potential benefits of mesothelin targeted cFP for NSCLC patients retaining the ability to consent and those who lose capacity to consent during the course of the trial outweigh the risks associated with the proposed entry-into-human trial with LMB-100 + pembrolizumab
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11.6 consent process and documentation

The informed consent document will be provided as a physical or electronic document to the participant or consent designee(s) as applicable for review prior to consenting. A designated study investigator will carefully explain the procedures and tests involved in this study, and the associated risks, discomforts and benefits. In order to minimize potential coercion, as much time as is needed to review the document will be given, including an opportunity to discuss it with friends, family members and/or other advisors, and to ask questions of any designated study investigator. A signed informed consent document will be obtained prior to entry onto the study. The initial consent process as well as re-consent, when required, may take place in person or remotely (e.g., via telephone or other NIH approved remote platforms used in compliance with policy, including HRPP Policy 303) per discretion of the designated study investigator and with the agreement of the participant/consent designee(s). Whether in person or remote, the privacy of the subject will be maintained. Consenting investigators (and participant/consent designee, when in person) will be located in a private area (e.g., clinic consult room). When consent is conducted remotely, the participant/consent designee will be informed of the private nature of the discussion and will be encouraged to relocate to a more private setting if needed. Consent will be documented with required signatures on the physical document (which includes the printout of an electronic document sent to participant) or as described below, with a manual (non-electronic) signature on the electronic document. When required, witness signature will be obtained similarly as described for the investigator and participant.
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manual (non-electronic) signature on electronic

When a manual signature on an electronic document is used for the documentation of consent at the NIH Clinical Center, this study will use the following to obtain the required signatures:  
Adobe platform (which is not 21 CFR Part 11 compliant); or, iMedConsent platform (which is 21 CFR Part 11 compliant)
During the consent process, participants and investigators will view individual copies of the approved consent document on screens at their respective locations (if remote consent); the same screen may be used when in the same location but is not required. Both the investigator and the participant will sign the document using a finger, stylus or mouse. Note: Refer to the CCR SOP PM-2, Obtaining and Documenting the Informed Consent Process for additional information (e.g., verification of participant identity when obtaining consent remotely) found here.  
11.6.1 Consent Process for Adults Who Lack Capacity to Consent to Research Participation For participants addressed in section **11.3**, an LAR will be identified consistent with Policy 403 and informed consent obtained from the LAR, as described in Section **11.6**.
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12 regulatory and operational considerations

This study may be temporarily suspended or prematurely terminated if there is sufficient reasonable cause. Written notification, documenting the reason for study suspension or termination, will be provided by the suspending or terminating party to study participants, IND sponsor and regulatory authorities. If the study is prematurely terminated or suspended, the Principal Investigator (PI) will promptly inform study participants, the Institutional Review Board (IRB), and sponsor and will provide the reason(s) for the termination or suspension. Study participants will be contacted, as applicable, and be informed of changes to study visit schedule. Circumstances that may warrant termination or suspension include, but are not limited to:  
Determination of unexpected, significant, or unacceptable risk to participants Demonstration of efficacy that would warrant stopping Insufficient compliance to protocol requirements Data that are not sufficiently complete and/or evaluable Determination that the primary endpoint has been met Determination of futility Study may resume once concerns about safety, protocol compliance, and data quality are addressed, and satisfy the sponsor, IRB and as applicable, Food and Drug Administration (FDA).
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12.2 quality assurance and quality

The clinical site will perform internal quality management of study conduct, data and biological specimen collection, documentation and completion. An individualized quality management plan will be developed to describe a site's quality management. Quality control (QC) procedures will be implemented beginning with the data entry system and data QC checks that will be run on the database will be generated. Any missing data or data anomalies will be communicated to the site(s) for clarification/resolution. Following written Standard Operating Procedures (SOPs), the monitors will verify that the clinical trial is conducted and data are generated and biological specimens are collected, documented (recorded), and reported in compliance with the protocol, International Conference on Harmonisation Good Clinical Practice (ICH GCP), and applicable regulatory requirements (e.g., Good Laboratory Practices (GLP), Good Manufacturing Practices (GMP)). The investigational site will provide direct access to all trial related sites, source data/documents, and reports for the purpose of monitoring and auditing by the sponsor, and inspection by local and regulatory authorities.
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12.3 conflict of interest policy

The independence of this study from any actual or perceived influence, such as by the pharmaceutical industry, is critical. Therefore, any actual conflict of interest of persons who have a role in the design, conduct, analysis, publication, or any aspect of this trial will be disclosed and managed. Furthermore, persons who have a perceived conflict of interest will be required to have such conflicts managed in a way that is appropriate to their participation in the design and conduct of this trial. The study leadership in conjunction with the National Cancer Institute has established policies and procedures for all study group members to disclose all conflicts of interest and will establish a mechanism for the management of all reported dualities of interest.
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12.4 confidentiality and privacy

Participant confidentiality and privacy is strictly held in trust by the participating investigators, their staff, and the sponsor(s). This confidentiality is extended to cover testing of biological samples and genetic tests in addition to the clinical information relating to participants. Therefore, the study protocol, documentation, data, and all other information generated will be held in strict confidence. No information concerning the study or the data will be released to any unauthorized third party without prior written approval of the sponsor. All research activities will be conducted in as private a setting as possible. The study monitor, other authorized representatives of the sponsor, representatives of the Institutional Review Board (IRB), and/or regulatory agencies may inspect all documents and records required to be maintained by the investigator, including but not limited to, medical records (office, clinic, or hospital) and pharmacy records for the participants in this study. The clinical study site will permit access to such records. The study participant's contact information will be securely stored at the clinical site for internal use during the study. At the end of the study, all records will continue to be kept in a secure location for as long a period as dictated by the reviewing IRB, Institutional policies, or sponsor requirements. Study participant research data, which is for purposes of statistical analysis and scientific reporting, will be stored at the NCI CCR. This will not include the participant's contact or identifying information. Rather, individual participants and their research data will be identified by a unique study identification number. The study data entry and study management systems used by clinical sites and by NCI CCR research staff will be secured and password protected. At the end of the study, all study databases will be archived at the NCI CCR. To further protect the privacy of study participants, a Certificate of Confidentiality has been issued by the National Institutes of Health (NIH). This certificate protects identifiable research information from forced disclosure. It allows the investigator and others who have access to research records to refuse to disclose identifying information on research participation in any civil, criminal, administrative, legislative, or other proceeding, whether at the federal, state, or local level. By protecting researchers and institutions from being compelled to disclose information that would identify research participants, Certificates of Confidentiality help achieve the research objectives and promote participation in studies by helping assure confidentiality and privacy to participants.
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13 pharmaceutical and investigational device

In order to be eligible for the study, participants are required to have positive mesothelin expression in archival tumor tissue, defined as at least 25% of tumor cells expressing mesothelin. Mesothelin expression testing is not FDA approved for this purpose; however, it is being used as an *in-vitro* diagnostic device. Validation assays to support the use of the IVD assay have been submitted to IND. All of the documentation is in the IND files. According to 21 CFR 812.3(m), a significant risk device presents a potential for serious risk to the health, safety and welfare of a subject and meets the significant risk criteria listed in the table below along with the sponsor's conclusions with regard to the applicability of these criteria to the current study. The device has been assessed by the sponsor as non-significant risk per the below.  
| current study. The device has been assessed by the sponsor as non-significant risk per the below.  Applicable  to current  Justification study Is an implant No The mesothelin test is not introduced into the subject  Is used in supporting or  sustaining human life No The device is diagnostic  While the device is diagnostic, we do not believe it  presents a potential for serious risk to the health and  welfare of the subject. The assessment of mesothelin  positivity is only used in tumors that may not have a  ubiquitous mesothelin expression and is assessed to help  to increase the possibility that all persons enrolling on  the study might derive benefit from therapy. Persons  that are deemed ineligible to enroll on the basis of this  test are eligible for studies within TGMB that are not  reliant on this test.  Is of substantial  importance in diagnosing  mitigating or treating  disease or preventing  impairment of human  health  No  Testing will be performed on archival samples or on  fresh tissue that is collected at screening for  confirmation of diagnosis. No additional collection of  tissue will occur for purposes of mesothelin testing.  Source: Testing will be performed via an IHC assay developed by the NCI Laboratory of  Pathology.  Otherwise poses a risk No   |
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