Purpose

Define user requirements for a standalone FUZKK utility that accepts laboratory CSV data, evaluates the records using limits embedded in code, and produces LabWare-compatible JSON.

Scope

The utility is intended for preliminary QC/QA review, integration testing, LIMS/LabWare flow and evidence-trail preparation. Final release decisions remain under the laboratory's validated procedure and responsible personnel.

Users

QC analyst, QA reviewer, CSV/validation engineer, LIMS/LabWare integration engineer, responsible laboratory specialist.

User requirements

1. The utility shall run without arguments and print its self-description, a built-in input.csv example from GetDemoData(), and demo evaluation for the embedded records.
2. The utility shall run with two arguments: input.csv output.json.
3. The utility shall not read input.csv and shall not write output.json when started without arguments.
4. CSV numeric values shall be parsed using CultureInfo.InvariantCulture.
5. Output shall be generated as LabWare-compatible JSON with Header, Samples, Results, Status, StatusCode, ErrorMessage, Description and DescriptionEN.
6. For PASS records, ErrorMessage shall be an empty string.
7. Embedded limits shall follow this priority: Ph. Eur. → British Pharmacopoeia / UK implementation → EAEU / regional requirements → EMA/ICH/EU guidance → USP fallback.
8. If an exact monograph is not known, strict standard API limits are used where applicable: assay 98–102%, total impurities ≤1.0%, individual impurity ≤0.5%.
9. For biologics and mAb-like products, aggregation, sterility and endotoxin checks shall be included where relevant to the utility purpose.
10. If a parameter may arrive in different units, the unit shall be represented as a separate input field or explicitly reflected in the input.csv field name.

Input CSV

BatchNumber,ADCName,ToxinType,FreePayload_ng_ml,DegradedPayload_ng_ml,LinkerDrug_ng_ml,TotalRelatedSubstances_ng_ml,LOD_ng_ml,LOQ_ng_ml,StabilityDays
ADC-FP-2026-001,Trastuzumab-Deruxtecan,DXd,15.0,5.0,10.0,30.0,1.0,3.0,30
ADC-FP-2026-002,Sacituzumab-Govitecan,SN-38,25.0,8.0,12.0,45.0,1.5,4.0,15
ADC-FP-2026-003,Polatuzumab-Vedotin,MMAE,5.0,1.0,2.0,8.0,0.5,1.5,60

input.csv fields

Utility description

ADC Free Payload Quantifier — Quantitative analysis of free toxin in ADC

ADC Free Payload Quantifier — Quantitative analysis of free toxin in ADC

ℹ️ Utility controls levels of free toxin and its degradation products:
• Free toxin (≤50 ng/mL)
• Degradation products (≤50 ng/mL)
• Linker-drug (≤50 ng/mL)
• Total related substances (≤100 ng/mL)

⚠️ CRITICAL: Free toxin causes systemic toxicity!
For MMAE/DXd limits are extremely low due to high potency.

Usage:
ADCFreePayloadQuantifier.exe → demo mode (console output)
ADCFreePayloadQuantifier.exe input.csv output.json → evaluate your data

Input format:
BatchNumber,ADCName,ToxinType,FreePayload_ng_ml,DegradedPayload_ng_ml,LinkerDrug_ng_ml,TotalRelatedSubstances_ng_ml,LOD_ng_ml,LOQ_ng_ml,StabilityDays

Example:
ADC-FP-2026-001,Trastuzumab-Deruxtecan,DXd,15.0,5.0,10.0,30.0,1.0,3.0,30

— WHY IS THIS NEEDED?
Free toxin control is critical for ADC safety (Hengrui, BeiGene, Hansoh):
• Free toxin (unbound to antibody) circulates throughout body, causing severe side effects
• MMAE causes neurotoxicity and neutropenia
• DXd can cause interstitial lung disease (ILD)
• Linker degradation products may also be toxic
• Regulators (FDA/NMPA) require ultra-sensitive control methods (LC-MS/MS)

⚠️ CRITICAL:
• Free toxin ≤50 ng/mL — strict limit for highly toxic warheads
• Degradation products ≤50 ng/mL — linker stability control
• Linker-drug ≤50 ng/mL — intermediate degradation product
• Total related substances ≤100 ng/mL — overall purity from free forms
• Method must have low limit of detection (LOD < 1 ng/mL)

Key features:
• Separate accounting of free toxin, degradation products, and linker-drug
• Control of total load of free forms
• Method sensitivity assessment (LOD/LOQ)
• Monitoring of impurity accumulation during storage (stability)

Critical parameters:
• Free toxin: ≤50 ng/mL
• Degradation products: ≤50 ng/mL
• Linker-drug: ≤50 ng/mL
• Total related substances: ≤100 ng/mL

💡 Usage tips:
1. Use LC-MS/MS with isotope-labeled standards for accuracy
2. Toxin extraction from sample must be efficient (liquid-liquid extraction)
3. Control toxin stability in solution during sample preparation
4. Compare free toxin levels at beginning and end of shelf life
5. If limits exceeded, batch rejected due to unacceptable toxicity risk for patient

⚠️ Note: Unlike standard impurities, free toxin in ADC is a highly active pharmacological agent. Even trace amounts (ng/mL) can have clinical significance. Chinese developers pay special attention to purification of drug from free toxin at dialysis or tangential flow filtration (TFF) stage.

Traceability and limitations

• The URS is used as the source document for functional specification, CSV review and later validation work.
• This document does not replace an approved pharmacopoeial monograph, validated analytical method or internal product specification.
• For product-specific limits, the approved customer specification takes priority.

The functional specification describes the behaviour of the standalone FUZKK console utility, input-data format, evaluation algorithm and output JSON structure.

Functional flow

1. Main() checks the number of arguments.
2. If no arguments are provided: PrintHello() prints the description and built-in input.csv example, then RunDemoEvaluation() executes Evaluate() over GetDemoData() and prints demo JSON.
3. If two arguments are provided: RunWithFiles(input.csv, output.json) reads CSV, evaluates each record and writes LabWare-compatible JSON.
4. LoadData() uses CultureInfo.InvariantCulture and shall not be called in no-arguments mode.
5. Evaluate() returns a named tuple with BatchNumber, ProductName, Parameters, CriticalFailCount, WarningCount, Recommendation and RecommendationEN.
6. GetIssues() builds messages for ErrorMessage in WARNING/FAIL cases.

Evaluation rules

• PASS: CriticalFailCount = 0 and WarningCount = 0.
• WARNING: CriticalFailCount = 0 and WarningCount > 0.
• FAIL: CriticalFailCount > 0.
• ERROR: exception during reading or processing.
• ErrorMessage remains empty for PASS.
• Limits are embedded in Program.cs; no external limit configuration is required.

Input and fields

BatchNumber,ADCName,ToxinType,FreePayload_ng_ml,DegradedPayload_ng_ml,LinkerDrug_ng_ml,TotalRelatedSubstances_ng_ml,LOD_ng_ml,LOQ_ng_ml,StabilityDays
ADC-FP-2026-001,Trastuzumab-Deruxtecan,DXd,15.0,5.0,10.0,30.0,1.0,3.0,30
ADC-FP-2026-002,Sacituzumab-Govitecan,SN-38,25.0,8.0,12.0,45.0,1.5,4.0,15
ADC-FP-2026-003,Polatuzumab-Vedotin,MMAE,5.0,1.0,2.0,8.0,0.5,1.5,60

Output JSON

{
  "Header": {
    "UtilityName": "ADC_FreePayload_Quantifier",
    "Version": "1.0.0",
    "Timestamp": "UTC",
    "InstrumentID": "FUZKK-QC-WORKSTATION",
    "OperatorID": "Admin"
  },
  "Samples": [
    {
      "SampleID": "from BatchNumber",
      "BatchNumber": "from CSV",
      "ProductName": "from CSV",
      "TestName": "utility-specific test",
      "AnalysisCode": "utility-specific code",
      "Status": "PASS | WARNING | FAIL | ERROR",
      "StatusCode": "1 | 2 | 0 | -1",
      "ErrorMessage": "",
      "Description": "Russian recommendation",
      "DescriptionEN": "English recommendation",
      "Results": [
        {
          "ParameterName": "parameter",
          "ResultValue": 0.0,
          "UnitOfMeasure": "unit",
          "SpecificationLimit": "limit",
          "IsWithinSpec": true
        }
      ]
    }
  ]
}