IMPD preparation example 3: [177Lu]DOTA-peptide
The present template has been drafted with the aim to provide a suitable “track” in case of preparation of a useful therapeutic radiopharmaceutical. 177Lu has been selected, among other suitable therapeutic radionuclides, due to its particularly favourable and well known properties, but it has to be clear that other radionuclides might be selected as well. As for the ligand, a generic “peptide” has been chosen, instead of a specific molecule, due to the high number of possible radionuclide/chelator/ligand complexes which have, on the other hand, a common preparation pathway. A generic 177Lu labelled DOTA-peptide was selected since DOTA is one of the most popular and commonly used chelators with high affinity for the radionuclide. As a consequence, most of the following sections do not contain real, experimental data, as shown with the other two templates dedicated to [11C]choline and [68Ga]DOTA-NOC. However, it is the opinion of the authors of the present document that the information included hereis a useful guide to the preparation of an IMPD for similar radiopharmaceuticals. Indeed [177Lu]DOTA-peptides, as well as other suitable therapeutic radiopharmaceuticals, may be prepared using different radionuclide sources, using automated system or even manually, and the automated system may be based on cassettes or not. The template presented here has been prepared hypothesizing the use of a “common” automated module. The proposed example cannot account, for obvious reasons, for all of the above mentioned methods and techniques, and it includes information related to one of the possible radionuclide/chelator/peptide preparation routes.
It is of paramount importance to remember that, in the preparation of the IMPD, every applicant should include the specific description of their own instrumentation, radiosynthetic pathway, experimental conditions, purification methods, data, etc. and also define their specifications with an appropriate justification.
2.2.1.S DRUG SUBSTANCE
2.2.1.S.1.1 Nomenclature
Drug substance
177Lu-(1,4,7,10-tetraazacyclododecane-N,N,N,N-tetraaceticacid)-peptide; [177Lu]Lu-DOTA-peptide
Radionuclide: Lutetium-177
2.2.1.S.1.2 Structure
Fig. 1– Structure of [177Lu]DOTA-peptide
Molecular formula: CxHy177LuNwOzS
Molecular weight of peptide: _____
2.2.1.S.1.3 General Properties
Physico-chemical properties:
Description of [177Lu]DOTA-peptide (any relevant information should be briefly stated, e.g. receptors affinity, therapeutic indications, etc.).
Nuclear properties of 177Lu are summarized as follows (Table 1):
Half-life: / 6.734daysPrincipal Photons: / 0.208 (11%), 0.113 (6.4%) MeV
Maximum Beta Energy: / 0.497 MeV (79%)
MaximumRange of Beta in Air: / 1.35 m
MaximumRange of Beta in Water: / 1.6 mm
Half-Value Layer for Lead: / 0.6 mm
Table 1 – Physical properties of 177Lu(http://ie.lbl.gov/toi/nuclide.asp?iZA=710177)
Tables 2 and 3 list representative physicochemical characteristics of the precursor DOTA-peptide and the radionuclide.
Physicochemical characteristics of DOTA-peptideAppearance / Related information of DOTA-peptide
Reconstitution / Related information of DOTA-peptide
pH / Related information of DOTA-peptide
LD50 / Related information of DOTA-peptide
Table 2 – List of selected physicochemical properties
177Lu specifications (obtained from the producer)Appearance / Colorless, clear solution
177mLu (HPGe)
All other impurities (HPGe) / Less than 0,05%
as of production date
Less than 0,01%
as of production date
Non-radioactive metal contamination (ICP) / Iron: < 20 µg/Ci
Zinc: < 40 µg/Ci
Aluminum: < 30 µg/Ci
Copper: < 30 µg/Ci
Calcium: < 100 µg/Ci
Lead: < 20 µg/Ci
Radionuclidic purity (HPGe) / ≥ 99%
pH / 1-3
Endotoxine level / <5 EU/mL
Table 3 – Characteristics of 177Lu
2.2.1.S.2.1 Manufacturer(s)
(To be filled in with data of DOTA-peptide supplier/manufacturer)
Institution nameAddress
Responsible person
e-mail address
Phone
Fax
(To be filled in with data of 177Lu supplier/manufacturer)
Institution nameAddress
Responsible person
e-mail address
Phone
Fax
(To be filled in with data of [177Lu]DOTA-peptide producer – small scale preparation institution)
InstitutionnameAddress
Person responsible for the small scale preparation of radiopharmaceuticals
e-mail address
Phone
Fax
Table 4 – Details of manufacturer(s)
2.2.1.S.2.2 Description of Manufacturing Process and Process Controls
A description from the supplier/manufacturer of the precursor should be added.
A description from the supplier/manufacturer of the 177Lu should be added.
[177Lu]DOTA-peptide is prepared from precursor DOTA-peptide and 177Lu chloride, which is withdrawn from activity vial and transferred to the reaction vial with corresponding amounts of peptide and labelling buffer. After the incubation at selected temperature, the desired product is purified at the end of the labelling reaction using a C18 cartridge which retains [177Lu]DOTA-peptide and releases free 177Lu. After washing the cartridge with physiological saline, the[177Lu]DOTA-peptide is eluted with a mixture of water/ethanol. The final product is then diluted with saline. The final product is sterilized by filtration through an inline 0.22 µm microbiological filter.
Due to the high emission energy of177Lu and multi-step synthesis, [177Lu]-DOTA-peptide is prepared using fully automated radiosynthesis modules. They are capable to perform all the necessary operations, from withdrawing 177Lu from the activity vial to the final formulation as an injectable solution of the radiopharmaceutical. For these reasons, the radiopharmaceutical preparations have to be considered as continuous processes carried out in closed systems. As a consequence, the active substance, as well as intermediates or by-products, are as a rule not isolated.
2.2.1.S.2.3 Control of Materials
A description regarding “Control of Materials” from the supplier/manufacturer of the precursor should be added.
A description regarding “Control of Materials” from the supplier/manufacturer of the 177Lu should be added.
The precursor DOTA-peptide should be manufactured according to GMP for API starting materials. As for the other starting materials, they should be preferably of pharmaceutical grade. In Table 5 a list of the starting materials used in the preparation of [177Lu]DOTA-peptide is reported.
Material / TestCassette (“kit”) / Visual inspection and control of each connection in aseptic condition (laminar flow cabinet class A)
DOTA-peptide / See the attached analysis certificate.
HCl / See the attached analysis certificate.
Ingredients for corresponding buffer / See the attached analysis certificate.
Water for injections / See the attached analysis certificate.
NaCl 0,9% injectable solution / See the attached analysis certificate
Ethanol / See the attached analysis certificate.
Any others / See the attached analysis certificate.
Table 5 – List of the starting materials used in the preparation of [177Lu]DOTA-peptide
2.2.1.S.2.4 Control of Critical Steps and Intermediates
The “in-process” controls are limited to the monitoring of the critical parameters (e.g. activity, reaction temperatures and pressures) through the graphical control software interface. Printouts of representative preparation process diagrams are usually provided. A full quality control program is set for the final product (see section # 2.2.1.P.4.2.)
2.2.1.S.2.5 Process Validation and/or Evaluation
Please refer to the section 2.2.1.P.3.5
2.2.1.S.2.6. Manufacturing Process Development
A description regarding “Manufacturing Process Development” from the supplier/manufacturer of the precursor should be added.
A description regarding “Manufacturing Process Development” from the supplier/manufacturer of the 177Lu should be added.
For [177Lu]DOTA-peptide please refer to the section 2.2.1.P.2.3
2.1.2.S.3 Characterization:
2.1.2.S.3.1 Elucidation of Structure and other Characteristics
A description regarding “Characterization” from the supplier/manufacturer of the precursor should be added.
Non radioactive Lu-DOTA-peptide and unchelated DOTA-peptide are used as references and are provided with a suitable Certificate of Analysis. HPLC analysis of reference standardsis performed using the same conditions as defined in the suitable European Pharmacopoeia monograph or the producer specifications in case a monograph does not exist. No further structure elucidating analyses are required. The experimental conditions are given in the section # 2.2.1.P.5.2.
2.1.2.S.3.2 Impurities
Radionuclidic purity:
A description regarding “Impurities” from the supplier/manufacturer of the 177Lu should be added.
Radiochemical purity
The analytical method for radiochemical purity determination is HPLC with integrated radioactivity detector. More details are given in section 2.2.1.P.5.2. The most plausible radiochemical impurity is Lu(III) ions as an unreacted species. The other potential radiochemical impurities are represented by radiolysis products.
Chemical purity
The most plausible chemical impurity in [177Lu]DOTA-peptide preparations is ethanol, which is used for radiosynthesis (solvent of class C).
As for the residual solvents, their limits are defined in the document “EMEA note for guidance on impurities: residual solvents” (CPMP/ICH/283/95), and inChapter 5.4 of Ph. Eur. The analysis is usually performed using gas-chromatography. More details are given in the section 2.2.1.P.5.2.
2.2.1.S.4 Control of the Drug Substance:
A description regarding “Control of the Drug Substance” from the supplier/manufacturer of the precursor should be added.
Details on methods for [177Lu]DOTA-peptide analysis, their validation, the batch analysis, and the justification of specifications will be provided in the appropriate 2.1.P sub-sections.
2.2.1.S.5 Reference Standards or Materials:
A description regarding “Reference Standards or Materials” from the supplier/manufacturer of the 177Lu should be added.
The list of the reference standard is provided in Table 6. There are two distinct kind of reference standard: i) chemical standard, ii) radionuclide calibrated sources.
The chemical reference standards are commercially available, chemical grade products. The specifications for purity have been set by the supplier, and accepted by the applicant. The specifications and most of the analytical tests (e.g. HPLC, MS) are performed by the supplier, and described in the attached Certificate of Analysis. The chemical reference standards are re-tested for chemical purity by the applicant using HPLC, as detailed in the section 2.2.1.P.5.
The radioactive reference standards are used to verify the calibration status of the gamma spectrometer and the dose calibrator, respectively. Their composition and the activity(ies) of the radionuclide(s) at reference time and date are described in the attached Certificate of Analysis. The calibrated sources are metrologically referable to recognized standards, and a verification of their identity or purity is in this case not applicable.
Reference standard / Aim / Test / Acceptance criteriaDOTA-PEPTIDE / DOTA-PEPTIDE Identity / HPLC / Purity> 95%
Mononuclide source / Dose calibrator / Not applicable / Not applicabile
Table 6 - List of reference standards
2.2.1.S.6 Container Closure System
For the reasons stated above, container closure system for the[177Lu]DOTA-peptide will be described for the drug product, in the appropriate sections.
2.2.1.S.7 Stability
A description regarding “Stability” from the supplier/manufacturer of the DOTA-peptide should be added.
A description regarding “Stability” from the supplier/manufacturer of the 177Lu should be added.
2.2.1.P INVESTIGATIONAL MEDICINAL PRODUCT UNDER TEST
2.2.1.P.1 Description and Composition of the Investigational Medicinal Product:
Each vial contains X mL of a mixture physiologicalsaline/ethanol (9/1) solution of [177Lu]DOTA-peptide, with radioactive concentration in the range X - Y MBq/mL at reference time. The [177Lu]DOTA-peptide solution is contained in X mL borosilicate glass vials (Ph. Eur. type I), sealed with chlorobutyl rubber stopper (Ph. Eur. compliant), which is in turn secured with an aluminum flip-off cap.
Composition
The composition of the finished drug product is given in Table 7:
Component / Quantity / Function / Reference[177Lu]DOTA-peptide / X - Y MBq/mL / Active substance / N.A.
NaCl 0,9% solution / X mL / Excipient / Ph. Eur.
Ethanol absolute / X mL / Excipient / Ph. Eur.
Water for injection / X mL / Excipient / Ph. Eur.
Table 7 – Composition of the injectable solution of [177Lu]DOTA-peptide
2.2.1.P.2 Pharmaceutical Development
[177Lu]DOTA-peptide is prepared using an automated synthesis module, where 177Luchloride is withdrawn from the activity vial and transferred to the reaction vial with the corresponding amount of peptide and labelling buffer. After the incubation at appropriate temperature the desired product is purified using a C18 cartridge which retains [177Lu]DOTA-peptide and releases free 177Lu. After washing the cartridge with physiological saline, [177Lu]DOTA-peptide is eluted with a mixture of water/ethanol.
The final product is then diluted with physiological saline to a final volume of X mL and sterilized by filtration through an inline 0,22 µm microbiological filter. The active substance is not isolated in the course of the preparation process, and it is available for quality control only at the end of the formulation step.
Stability studies have been performed, demonstrating that the composition of the [177Lu]DOTA-peptide solution does not undergo significant changes to chemical and radiochemical purity during the assigned shelf life (e.g. 24 h). For more information about stability, see section 2.2.1.P.8.
2.2.1.P.2.3 Manufacturing Process Development
Not applicable
2.2.1.P.3 Manufacture:
2.2.1.P.3.1 Manufacturer(s)
InstitutionNameAddress
Person responsible for the small scale preparation of radiopharmaceuticals
e-mail address
Phone
Fax
2.2.1.P.3.2 Batch Formula
A batch of [177Lu]DOTA-peptide usually consists of a single, multi-dose vial. The materials used in the preparation of a typical batch using automated module are listed in Table 8.
Starting materials / AmountsDOTA-peptide / X µg
Sterile water (appropriate quality) / X mL
NaCl 0,9% physiological solution / X mL
Ethanol absolute / X mL
Labelling buffer / X mL
Specific sterile, single use, cassette / 1
Table 8 – List of starting materials used in a typical [177Lu]DOTA-peptide batch production
Batches of [177Lu]-DOTA-peptide to be used in clinical trials typically consist of a single vial.
2.2.1.P.3.3 Description of Manufacturing Process and Process Controls
The 177Lu-DOTA-peptide preparation process has already been described in section 2.2.1.S.2.2 of the present document. However, a flow chart is added below:
Fig. 2– Flowchart of the preparation process of [177Lu]DOTA-peptide
2.2.1.P.3.4 Controls of Critical Steps and Intermediates
Description of in-process controls and intermediated has already been given in section 2.2.1.S.2.4
2.2.1.P.3.5 Process Validation and/or Evaluation
Process validation has been performed by preparing and testing three consecutive batches of [177Lu]DOTA-peptide. Validation runs have been performed in the same operating conditions (e.g. starting 177LuCl3 activity, amount of precursor, reaction parameters) using the same instrumentation and starting materials in the stated quantities normally set for typical runs.
The parameters evaluated, and their acceptance criteria are reported in Table 9:
Parameters / Acceptance criteriaRadioactive concentration / X – Y MBq/mL
Final volume / X Y mL
Table 9 – Production parameters evaluated during the process validation
The experimental data are reported in the following table:
BATCH 1 / BATCH 2 / BATCH 3[177Lu]DOTA-peptide activity / ___ MBq / ___ MBq / ___ MBq
Radioactive concentration / ___ MBq/mL / ___ MBq/mL / ___ MBq/mL
Volume / ___ mL / ___ mL / ___mL
Conform / □ yes□ no / □ yes□ no / □ yes□ no
Table 10 – Experimental production data for process validation
Each batch has been analyzed following the complete quality control program. The results are summarized in Table 11. Specifications are described in section 2.2.1.P.5.1.
QUALITY CONTROLBatch 1 / Batch 2 / Batch 3
Test / Acceptancecriteria / results / Conform / results / Conform / results / Conform
pH / 4.5-8.5 / ___ / □ Yes □ No / ___ / □ Yes □ No / ___ / □ Yes □ No
Appearance / Description of solution (i.e. clear solution) / ___ / □ Yes □ No / ___ / □ Yes □ No / ___ / □ Yes □ No
EtOH / ≤ 400 mg/V* / ___ / □ Yes □ No / ___ / □ Yes □ No / ___ / □ Yes □ No
DOTA-peptide / X - Y µg/mL / ___ / □ Yes □ No / ___ / □ Yes □ No / ___ / □ Yes □ No
Radiochemicalpurity / ≥ X% of [177Lu]DOTA-peptide / ___ / □ Yes □ No / ___ / □ Yes □ No / ___ / □ Yes □ No
Radionuclidic purity / ≥ 99% / See the attached analysis certificate / □ Yes □ No / See the attached analysis certificate / □ Yes □ No / See the attached analysis certificate / □ Yes □ No
Half-life / 6.65 days / See the attached analysis certificate / □ Yes □ No / See the attached analysis certificate / □ Yes □ No / See the attached analysis certificate / □ Yes □ No
Filter integrity / Online Pressure test at 2 Bar / ___ / □ Yes □ No / ___ / □ Yes □ No / ___ / □ Yes □ No
Sterility / Ph. Eur. Conform / ___ / □ Yes □ No / ___ / □ Yes □ No / ___ / □ Yes □ No
Bacterial endotoxins / Ph. Eur.Conform (<17,5 EU/mL) / ___ / □ Yes □ No / ___ / □ Yes □ No / ___ / □ Yes □ No
Table 11 – Experimental quality control data for process validation
*V = maximum recommended dose (mL)
2.2.1.P.4 Control of Excipients:
2.2.1.P.4.1 Specifications
The excipients used in the preparation process of the finished investigational radiopharmaceutical should meet the specifications reported in Table 12.
Excipient / Specification0.9% NaCl / Ph. Eur.
Ethanol / Ph. Eur.
Table 12 – List of excipients
The 0.9% NaCl and ethanol solutions are approved following a check for packaging integrity, expiry and Certificate of Analysis. The appearance should also be verified. For the above reasons, for excipient control there is no need to describe analytical procedures, as well as their validation and justification of specification(s).
2.2.1.P.5 Control of the Investigational Medicinal Product:
2.2.1.P.5.1 Specifications
Each batch of [177Lu]DOTA-peptide is submitted to quality control, with the aim to evaluate chemical, radiochemical and microbiological purity of the finished product. The QC tests are summarized in Table 13.
Parameters / Test / SpecificationAppearance / Visual inspection / Description of solution (i.e. clear solution).
Identification / HPLC / The principal peak in the radiochromatogram obtained with the test solution of 177Lu-DOTA-peptide has approximately the same retention time as the principal peak in the chromatogram obtained with a reference solution of “cold” Lu-DOTA-peptide.
Radiochemical purity / HPLC / [177Lu]DOTA-peptide ≥ X%
Residual solvent* / GC / Ethanol ≤ 400 mg/max. injectable volume
Radionuclidic purity / See the attached analysis certificate / 177Lu ≥ 99%
Half-life / See the attached analysis certificate / 6.65 days
pH / pH paper / X-Y
Bacterial endotoxins* / Ph. Eur. / <175 EU/max. injectable volume
Filter integrity / Bubble point / Following specification of the supplier
Sterility* / Ph. Eur. / Sterile
Table 13 – Specifications and acceptance criteria for [177Lu]DOTA-peptide
*Tests performed after the release of the radiopharmaceutical, due to both the test duration not compatible with radionuclide half-life (e.g. sterility test), and for radiation protection reasons (e.g. bacterial endotoxins test).
2.2.1.P.5.2 Analytical Procedures
Analytical procedures are described in details in relevant Standard Operating Procedures, which are available on request.
2.2.1.P.5.2.1 Identification and determination of the chemical and radiochemical purity using HPLC
Instrumentation:
HPLC (description of the instrument, detectors)
[177Lu]DOTA-peptide analysis: specifications
-Identification: the main radiochemical peak should have the same retention time evidenced by the SST test for standard Lu-DOTA-peptide ± 0.2 min
-Radiochemical purity: peak area for [177Lu]DOTA-peptide has to be > X % of thetotal peak areas
-Chemical purity (DOTA-peptide): the peak area for DOTA-peptide should not be more than the corresponding area of the peak obtained with reference SST solution
2.2.1.P.5.2.2 Determination of residual solvents using gas-chromatography